Estrogen receptor ligand treatment for neurodegenerative diseases

ABSTRACT

The present invention relates to treatment of neurological diseases such as multiple sclerosis (MS) and Alzheimer&#39;s disease, using an estrogen receptor beta (ERβ) ligand.

PRIORITY CLAIM

This application claims priority to U.S. Provisional Patent ApplicationNo. 62/044,679, filed on Sep. 2, 2014, and U.S. Provisional PatentApplication No. 62/047,285, filed on Sep. 8, 2014, each of which ishereby incorporated by reference in its entirety.

BACKGROUND

This invention relates generally to a novel treatment to preventneurodegcneration in the central nervous system due to diseases such asmultiple sclerosis (MS). Alzheimer's disease, Parkinson's disease,spinal cord injury, stroke, etc. More specifically, some embodiments ofthe present invention relates to a treatment comprising a combination ofan estrogen receptor ligand with a secondary agent, such as animmunotherapeutic compound.

GENERAL BACKGROUND

There are no neuroprotective drugs that can be taken for long durationsof time without significant side effects. Estrogens, as well as the useof estrogen receptor (ER) alpha ligand treatments, have been studied indisease and injury models and in humans. Estrogen, and estrogen receptoralpha ligand treatments, are effective in some disease and injurymodels. For example, they are both anti-inflammatory and neuroprotectivein experimental autoimmune encephalomyelitis (EAE), the animal model formultiple sclerosis (MS) and there is a dose response whereby higherlevels are more protective. However, in humans, treatment with estrogensor ER alpha ligands may not be tolerable due to the induction of breastcancer and uterine cancer, which are mediated by estrogen receptor alphain the breast and uterus, respectively. One must always consider therisk:benefit ratio of any estrogen treatment when considering its use inneurodegenerative diseases. Estrogens in the form of hormone replacementtherapy have been associated with side effects and therefore arecontroversial with respect to whether they are recommended for use in asubset of healthy menopausal women, depending on their menopausalsymptoms and other disease risk factors. While the risk:benefit ratio indebilitating neurodegenerative diseases is clearly different than therisk:benefit ratio in healthy individuals, optimizing efficacy andminimizing toxicity, remains the goal. Hence, determining which estrogenreceptor mediates the neuroprotective effect of estrogen treatment is ofcentral importance.

In contrast, estrogen receptor beta (ERP) is not associated with breastor uterine cancer. Thus, estrogen receptor beta ligands may be used forlong durations and/or for patients with some risk factors for breast anduterine cancer who could not otherwise tolerate estrogen or estrogenreceptor alpha ligand treatment.

One estrogen, estradiol, and estrogen receptor alpha ligands agonisthave been shown to be both anti-inflammatory and neuroprotective in theEAE model. They ameliorate EAE symptomology immediately after thedisease is induced. In contrast, estrogen receptor beta ligand treatmentis not anti-inflammatory, but has been shown for the first time to beneuroprotective. This mechanism is thought to explain why ER beta ligandtreatment does not work at EAE onset, but does work later to promoterecovery or delay EAE progression.

There are currently no purely neuroprotective treatments for MS. Thus,for diseases such as MS, which have both an inflammatory and aneurodegenerative component, estrogen receptor beta ligands may beuseful. For diseases that do not appear to have an inflammatorycomponent, but only a neurodegenerative component, then the estrogenreceptor beta ligand treatment may also be useful. Notably, Alzheimer'sdisease, Parkinson's disease, brain or spinal cord injury and stroke areprimarily purely neurodegenerative diseases or injuries, but there maybe a minor inflammatory component. To date, for Alzheimer's disease, forexample, there are only treatments that can be used in short termduration. Hence, the identification of an alternative neuroprotectiveagent represents an important advance in preclinical drug development inMS and other chronic neurodegenerative diseases or injuries.

SUMMARY

The present invention is directed to a treatment for neurodegenerationin the central nervous system due to diseases such as MS. Parkinson'sdisease, cerebellar ataxia, Down's Syndrome, epilepsy, strokes.Alzheimer's disease, as well as brain and/or spinal cord (CNS) injury.

In accordance with some embodiments of the present invention, a methodfor treating the symptoms of a neurodegenerative disease or CNS injuryin a mammal is provided, the method comprising the steps ofadministering to the mammal a primary agent being an estrogen receptorligand, and optionally, a secondary agent, such as an immunotherapeuticcompound. In one embodiment, the estrogen receptor ligand may be an ERβligand, such as AC-186 (Compound I) or a compound substantially similarin chemical structure and/or activity thereto (such as those compoundsdisclosed in PCT Patent Publication WO 2013/017619 A1, incorporated byreference herein).

In some embodiments, the estrogen receptor beta ligand is a compoundselected from the compounds disclosed in U.S. Patent ApplicationPublications 2012/0202861 A1 or 2013/0131061 A1 (incorporated herein byreference). In some embodiments, the estrogen receptor beta ligand isKBRV1 or KBRV2 (Karo Bio, Huddinge, Sweden).

In accordance with other embodiments of the present invention, theinvention comprises the use of a primary agent comprising an estrogenreceptor beta ligand for a neuroprotective effect. In certain embodiment% an ERβ ligand may be used to prevent and/or delay the onset orprogression of disease or injury after the acute phase and/or ameliorateclinical symptoms of neurodegenerative diseases or injury, includingmultiple sclerosis. In one embodiment, a secondary agent, including animmunotherapeutic or anti-inflammatory compound could be used. Incertain embodiments, the secondary agent is an immunotherapeutic agent,and the amount of the immunotherapeutic agent administered incombination with the estrogen receptor beta ligand is less than atherapeutically effective amount when the immunotherapeutic agent isadministered alone.

In accordance with some embodiments the present invention relates to useof at least one primary therapeutically active agent, the primarytherapeutically active agent being an estrogen receptor beta ligand,optionally in combination with a secondary active agent for themanufacture of a medicament for the therapeutic treatment of aneurodegenerative disease in a mammal.

The above described and many other features and attendant advantages ofthe present invention will become apparent from a consideration of thefollowing detailed description when considered in conjunction with theaccompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1. EAE scores versus time for all treatment groups in femaleC57BL/6 mice. The group receiving 30 mg/kg of AC-186 (“▾”) displayed thebest EAE score. Treatment with DPN was not effective when sesame oil wasthe carrier. This loss of effect of DPN with sesame oil as the carriersuggests that the carrier is important since previous work had shownthat DPN ameliorated EAE when provided in a different carrier (Miglyol).Despite using the sesame carrier, some effectiveness was observed forAC-186 at the highest dose (30 mg/kg).

FIG. 2. EAE scores separated by different doses of AC-186 dose. Eachgraph shows the scores for control animals (Veh; “●”). Animals thatreceived high dose AC-186 treatments (30 mg/kg; lower panel, “▾”) hadsignificantly less severe EAE scores as compared to the sesame oilvehicle alone (p=0.0299).

FIG. 3. Rotarod to assess motor coordination and balance. Rotarod timesfor female C57BL/6 mice that received high dose AC-186 treatments (30mg/kg; “▾”) in sesame oil versus control animals (Veh; “●”).

FIG. 4. Comparisons of outcomes with miglyol as the carrier, using DPNas the ER beta ligand. EAE scores and rotarod times for a positivecontrol DPN in the vehicle of 10% ethanol/miglyol solution (Migl. DPN;“▪”) and for control animals (Migl. Veh; “●”). DPN treated animalsdisplayed better EAE scores and better performance with the rotarodtest.

FIG. 5. Comparisons of outcomes using sesame oil versus miglyol as thecarrier, using AC-186 as the ER beta ligand (EAE scores). EAE scores formale C57BL/6 mice that received high dose AC-186 treatments (30 mg/kg)in either sesame oil (top and bottom graphs) or miglyol (top and middlegraphs) or the vehicle alone. Animals receiving AC-186 in miglyoldisplayed better EAE scores than those receiving the vehicle alone (see,e.g., middle panel).

FIG. 6. Comparisons of outcomes using sesame oil versus miglyol as thecarrier, using AC-186 as the ER beta ligand (Rotarod times). Rotarodtimes for male C57BL6 mice that received high dose AC-186 treatments (30mg/kg) in either sesame oil (top and bottom graphs) or miglyol (top andmiddle graphs) or the vehicle alone. Animals receiving AC-186 in miglyoldisplayed better rotarod scores than those receiving the vehicle alone(see, e.g., middle panel).

FIG. 7. Treatment with carrier alone and no ER beta ligand. EAE scoresfor animals that received either the sesame oil or miglyol vehicle andno ER ligand. There was no effect on disease, with either carrier, whenno ER beta ligand was administered, showing that the ER beta ligand'sdelivery in each carrier underlies differences in effectiveness witheach carrier.

FIG. 8 EAE scores in a different chronic progressive EAE model in NODfemales. EAE scores for NOD females under various treatment conditions.Animals receiving an estrogen receptor β ligand (either AC-186 at 30mg/kg, AC-186 at 10 mg/g, or DPN) in a miglyol vehicle each displayedbetter EAE scores than animals receiving vehicle alone. In contrast, noprotection was observed using the sesame oil carrier, even with a highdose AC-186 treatment.

FIG. 9. EAE Rotarod times for NOD females under various treatmentconditions. Rotarod performance was relatively poor in all female NODgroups with no significant difference between treatment groups.

FIG. 10. EAE scores in NOD males under various treatment conditions.Standard EAE disease scores were ameliorated with AC-186 at 30 mg/kg,but not with AC-186 at 10 mg/kg, dose, each as compared to carriervehicle alone (miglyol).

FIG. 11. EAE Rotarod times for NOD males under various treatmentconditions. Rotarod performance was relatively poor in all male NODgroups with no significant difference between treatment groups.

FIG. 12. EAE scores for C57BL/6 females under optimized treatmentconditions. Standard EAE disease scores were ameliorated with AC-186 at30 mg/kg and 10 mg/kg (p<0.0001), each as compared to vehicle (miglyol).

FIG. 13. Rotarod times for C57BL/6 females under optimized treatmentconditions. Rotarod testing showed an improvement with the AC-186 30mg/kg dose (p=0.0005).

FIG. 14. AC-186 post-treatment during EAE: effects on axonal densities,beta-APP, and myelin in spinal cord. Pathology was performed on spinalcords of C57BL/6 females with EAE under optimized treatment conditionswith AC-186 administered at either 30 mg/kg or 10 mg/kg, each in miglyolversus carrier vehicle alone. FIG. 14 consists of two panels, labeledpanels (A) and (B). (Panel A, top row) Representative 10× captures ofspinal cord sections at the dorsal column of matched healthy control(left), vehicle-treated EAE (2nd from left), AC-186 10 mg/kgpost-treated EAE (3rd from left), and AC-186 30 mg/kg post-treated EAE(right). EAE mice were sacrificed at EAE day 60. Axons and myelin werestained with NF200 and MBP, respectively (Panel A, top three rows).Representative 10× confocal images of spinal cord sections were stainedfor axonal damage using beta-APP (A, bottom row). (Panel B)Quantification of axonal densities (left), beta-APP expression (middle),and myelin staining intensity (right), Vehicle treated EAE (Veh) ascompared to matched healthy controls (Cont) showed significantly reducedaxon numbers (left), increased beta-APP (middle) and reduced myelin(right). AC-186 10 mg/kg, and 30 mg/kg treated EAE groups each showedsignificantly more axon numbers compared with the Vehicle treated EAEgroup (p<0.03, Veh vs AC-186 10 mg/kg and Veh vs AC-186 30 mg/kg). Thebeta-APP staining showed that the AC-186 30 mg/kg treatment group hadsignificantly less expression of beta-APP compared with the Vehicletreated EA E group (p=0.0296, Veh vs AC-186 30 mg/kg). The AC-186 10mg/kg treatment group showed a trend of less beta-API staining comparedwith the Vehicle treated EAE group, but the difference with this dosedid not reach significance. MBP staining showed that both AC-186treatment groups had a trend for somewhat higher MBP staining intensityas compared to vehicle but this did not reach statistical significance.Four mice in each treatment group were examined for each treatmentgroup, p-values were determined by one-way ANOVA.

FIG. 15. AC-186 post-treatment during EAE: effects on macrophages and Tcells. Further neuropathology was done on spinal cords from FIG. 14,here focusing on immune cell infiltrates. FIG. 15 consists of twopanels, labeled panels (A) and (B). (Panel A, top row) Representative10× captures of spinal cord sections at the dorsal column of matchedhealthy controls (left), vehicle-treated EAE (2nd from left), AC-186 10mg/kg post-treated EAE (3rd from left), and AC-186 30 mg/kg post-treatedEAE (right). EAE mice were sacrificed at EAE day 60. Immune cells in theCNS were stained with pan-leucocyte marker CD45. The tissues werecounterstained by DAPI. (Panel A, middle row) Representative 10×confocal images of spinal cord sections stained with Iba-1. (Panel A,bottom row) Representative 10× confocal images of spinal cord sectionsstained with CD3. (Panel B) Quantification of CD45 immunoreactivity(left), Iba-1 globoid to quantify macrophage like cells (middle), andCD3 to quantify T lymphocytes (right) as shown in (Panel A). Vehicletreated EAE (Veh) as compared to matched healthy controls (Cont) showedsignificantly increased CD45 staining (left), increased Iba-1 globoidcells (middle), and increased CD3 cells (right). AC-186 10 mg/kg and 30mg/kg post-treatment EAE groups each showed a reduction of CD45expression compared with the Vehicle treated EAE group (p=0.001, Veh vsAC-186 10 mg/kg, p=0.005, Veh vs AC-86 30 mg/kg). Each of the AC-186treatment groups also showed a significant reduction in the number ofIba-1 stained cells with globoid morphology as compared with the Vehicletreated EAE group (p<0.0001, Veh vs AC-186 10 mg/kg, p=0.0001, Veh vsAC-186 30 mg/kg). There were no differences in the number of CD3 stainedcells between either of the AC-186 treatment groups as compared to theVehicle EAE treatment group. Four mice in each treatment group (or threemice for Veh and AC-186 10 mg/kg groups in CD45 staining) were examinedfor each treatment group, p-values were determined by one-way ANOVA.

FIG. 16. EAE scores and rotarod times for female C57B/6 mice whoreceived 30 mg/kg AC-186 in miglyol for subsequent in vivo MRI of thebrain followed by neuropathologic analyses of cerebrum and cerebellum.(Top) Standard EAE disease scores were ameliorated in female C57BL′6mice treated with AC-186 at 30 mg/kg as compared to vehicle (miglyol),p<0.0001. (Bottom) Rotarod testing showed significant improvement withthe AC-186 30 mg/kg as compared to vehicle (miglyol), p<0.0001.

FIG. 17. AC-186 treatment protects against Whole Brain, Cortical andCerebellar Atrophy by in vivo MRI in EAE. FIG. 17 consists of threepanels, labeled panels (A), (B), and (C). (Panel A) A graph of the meanwhole brain volume in healthy controls (“X”), AC-186-treated mice withEA E (“O”), and vehicle-treated mice with EAE (“▪”) at d0, d30 and d60.AC-186-treated EAE mice exhibit less brain atrophy than vehicle-treatedEAE mice as early as d30. (Panel B) A graph of the mean cerebral cortexvolume in healthy controls, AC-186-treated mice with EAE andvehicle-treated mice with EAE at d0, d30 and d60. AC-186-treated EAEmice exhibit less atrophy in the cerebral cortex than vehicle-treatedEAE mice by d60. (Panel C) A graph of the mean cerebellar volume inhealthy controls, AC-186-treated mice with EAE and vehicle-treated micewith EAE at d0, d30 and d60. AC-186-treated EAE mice exhibit lesscerebellar atrophy than vehicle-treated EAE mice by d60.

FIG. 18. AC-186 treatment protects against loss of cerebral andcerebellar neurons and synapses in gray matter. (Top row) Quantificationof NeuN neuronal cells in the cerebral cortex gray matter (left), andpercent area of Post-Synaptic Density-95 (PSD-95, right) positivity inhealthy controls (Cont), Vehicle (Veh), and AC-186-treated EAE mice.Vehicle treated EAE mice, as compared to age-matched healthy controls,had fewer numbers of NeuN⁺ cortical neurons and less PSD-95 staining(p=0.02, NeuN; p=0.0328. PSD-95, one-way ANOVA). AC-186 treated EAEmice, as compared to Vehicle, had higher numbers of NeuN⁺ corticalneurons and greater PSD-95 staining, with values comparable toage-matched healthy controls (p=0.02, NeuN, p=0.0049, PSD-95, one-wayANOVA). (Bottom row) Quantification of Calbindin⁺ Purkinje cells (left)in the cercbellar gray matter, and PSD-95 (right) in cerebellar graymatter of age-matched healthy controls, Vehicle. and AC-186-treated EAEmice. Vehicle treated EAE mice, as compared to age-matched healthycontrols, had fewer numbers of Calbindin⁺ Purkinje cells and less PSD-95staining (p=0.02, Calbindin; p=0.0328, PSD-95; one-way ANOVA). AC-186treated EAE mice, as compared to Vehicle, had higher numbers ofCalbindin⁺ Purkinje cells and greater PSD-95 staining, with valuescomparable to age-matched healthy controls (p=0.04. Calbindin; p=0.0005,PSD-95; one-way ANOVA).

FIG. 19. AC-186 treatment protects against axonal loss, axonal damage,and myelin loss in spinal cord white matter. EA mice receiving AC-186 at30 mg/kg or vehicle were sacrificed. Quantification is shown of axonaldensities by NF200 staining (top), beta-APP expression for axonal damage(middle), and myelin staining intensity by MBP (bottom). The vehicletreated (Veh) group as compared to matched healthy controls (Cont)showed significantly reduced axon numbers (left), increased beta-APP(middle) and reduced myelin (right). The AC-8630 mg/kg treated EAE groupshowed significantly more axon numbers compared with the Vehicle treatedEAE group (p<0.05, Veh vs AC-186 30 mg/kg). The beta-APP staining showedthat the AC-186 30 mg/kg treated group had significantly less expressionof beta-APP compared with the Vehicle treated EAE group (p<0.04, Veh vsAC-196 30 mg/kg). MBP staining showed that the AC-186 treatment grouphad higher MBP staining intensity as compared to vehicle (p<0.004, Vehvs AC-186 30 mg/kg). Three to five mice were examined for each treatmentgroup, p-values were determined by one-way ANOVA. These results largelyreproduce the results in FIG. 14, showing a protective effect on axons.However, a beneficial effect of AC-186 as compared to vehicle withregard to increased MBP staining, which was observed as a trend in theexperiment in FIG. 14, was observed as significant in FIG. 19. Together,this data suggests that AC-186 treatment indeed has a beneficial effecton preserving not only axons, but also myelin, in the CNS.

FIG. 20. AC-186 treatment during EAE: effects on macrophages and Tcells. EAE mice receiving AC-186 at 30 mg/kg or vehicle were sacrificed.Quantification of CD45 immunoreactivity to quantify all immune cells(top), Iba-1 globoid to quantify macrophage like cells (middle), and CD3to quantify T lymphocytes (bottom) was done. Vehicle treated EAE (Veh)as compared to matched healthy controls (Cont) showed significantlyincreased CD45 staining (top), increased Iba-1 globoid cells (middle),and increased CD3 cells (bottom). AC-186 30 mg/kg treated EA E groupsshowed a reduction of CD45 expression compared with the Vehicle treatedEAE group (p=0.029, Veh vs AC-186 30 mg/kg). The AC-186 treatment groupalso showed a significant reduction in the number of Iba-1 stained cellswith globoid morphology as compared with the Vehicle treated EA E group(p=0.037, Veh vs AC-186 30 mg/kg). There were no differences in thenumber of CD3 stained cells between the AC-186 treated group as comparedto the Vehicle EAE treatment group. Three to five mice were examined foreach treatment group, p-values were determined by one-way ANOVA. Theseresults largely reproduce the results of FIG. 15, showing protectiveeffects on macrophages but not T cells.

FIG. 21. AC-186 treatment during EAE: effects on cerebellar whitematter. EAE mice receiving AC-186 at 30 mg/kg or vehicle weresacrificed. Quantification is shown of nyelin staining intensity by MBP(left) and axonal densities by NF200 staining (right). The vehicletreated (Veh) group as compared to matched healthy controls (Cont)showed significantly reduced myelin (left) and reduced axon numbers(right). MBP staining showed that the AC-186 treated group had higherMBP staining intensity as compared to vehicle (p<0.0176, Veh vs AC-18630 mg/kg). The AC-186 30 mg/kg treated EAE group showed significantlymore axon numbers compared with the Vehicle treated EAE group (p<0.01,Veh vs AC-186 30 mg/kg). Three to five mice were examined for eachtreatment group, p-values were determined by one-way ANOVA.

FIG. 22. AC-186 treatment during EAE: effects on cerebral white matter.Quantification is shown of myelin staining intensity by MBP (left) andaxonal densities by NF200 staining (right) in the splenium of the corpuscallosum of the cerebrum. The vehicle treated (Veh) group as compared tomatched healthy controls (Cont) showed a trend for reduced myelin, butthis did not reach significance (left), and a significant reduction inaxon numbers (p=0.0025, Cont vs Veh) (right). Regarding the effect ofAC-186 treatment, MBP staining showed that the AC-186 treated group hada trend for higher MBP staining intensity as compared to vehicle (left),but this did not reach significance. The AC-186 30 mg/kg treated EAEgroup showed significantly more axon numbers compared with the Vehicletreated EAE group (p=0.0286. Veh vs AC-186 30 mg/kg) (right). Three tofive mice were examined for each treatment group, p-values weredetermined by one-way ANOVA.

FIG. 23. AC-186 treatment during EAE: effects on cerebellar gray matterneurons and synapses. Quantification of Calbindin⁺ cells in the Purkinjecell layer, as well as PSD-95 and Synapsin-1 positivity in cerebellarmolecular layer of age-matched healthy controls, Vehicle, andAC-6-treated EAE mice. Vehicle treated EAE mice, as compared toage-matched healthy controls, had fewer numbers of Calbindin⁺ cells inthe cerebellar Purkinje cell layer and less PSD-95 and Synapsin1staining (p=0.02, Calbindin; p=0.0328, PSD-95; p=0.0021, Synapsin1,one-way ANOVA) in the molecular layer. AC-186 treated EAE mice, ascompared to Vehicle, had higher numbers of Calbindin® cells in thePurkinje cell layer and greater PSD-95 and Synapsin1 staining in themolecular layer, with values comparable to age-matched healthy controls(p=0.04, Calbindin: p=0.0005. PSD-95: p<0.0025, Synapsin1, one-wayANOVA).

FIG. 24. AC-186 treatment during EAE: effects on cerebral gray matterneurons and synapses. Quantification of NeuN neuronal cells, percentarea of Post-Synaptic Density-95 (PSD-95) and presynaptic Synapsin-1positivity in cerebral cortex of healthy controls (Cont), Vehicle (Veh),and AC-186-treated EAE mice. Vehicle treated EAE mice, as compared toage-matched healthy controls, had fewer numbers of NeuN⁺ corticalneurons and less PSD-95 staining in the cerebral cortex (p=0.02, NeuN;p=0.0328, PSD-95, one-way ANOVA). AC-186 treated EAE mice, as comparedto Vehicle, had higher numbers of NeuN⁺ cortical neurons and greaterPSD-95 staining in the cerebral cortex, with values comparable toage-matched healthy controls (p=0.02, NeuN, p=0.0049. PSD-95, one-wayANOVA). There were no significant differences in Synapsin-1 staining.Three to five mice were examined for each treatment group, p-values weredetermined by one-way ANOVA.

FIG. 25. Peripheral estrogens improve performance on the Morris watermaze via ERP expressed on neurons. Healthy female C57BL/6 (with no EAEinduced) were tested for cognitive performance (spatial memory) usingthe Morris water maze, whereby mice learn to spend more time in a targetquadrant (TQ) as compared to three other quadrants (OQ). (Top graph)Sham-Placebo mice (SP, n=6) spent more time in the target quadrant dueto proper learning, whereas ovariectomized, placebo-treated mice(OVX-Placebo, OP, n=7) did not show learning, thereby confirming thatthe loss of endogenous ovarian hormones causes cognitive disability. Incontrast, ovariectomized, estriol-treated mice (OVX-Estriol, OE, n=7)displayed that the ability to learn may be restored with estrioltreatment (**p≤0.01, ANOVA-Sidak's posttest). (Bottom graph) Todetermine (a) whether ER beta is required for proper learning and, morespecifically, (b) whether ER beta expression in neurons is required forlearning, mice were generated that had ER beta conditionally knocked outusing CreLox technology, with Cre driving the knock out in neurons viathe neuron-specific enolase (NSE) promoter. Wild-type SP (n=7), but notSP NSE-Crc:Erβ^(fl/fl) (n=7), OP NSE-Cre:Erβ^(fl/fl) was (n=7), or OENSE-Cre:Erβ^(fl/fl) (n=7) mice, spent significantly greater % durationin the TQ than in the OQ, averaged. (***p≤0.001, ANOVA-Sidak'sposttest). These results suggest that, when ER beta is removed fromneurons in the brain, mice lose the ability to learn. Further, thesefindings suggest that the ability of estriol to rescue learning ismediated by ER beta expression in neurons. The dashed line on the graphsindicates the time expected to be spent in a quadrant by random chancein the absence of learning (25%).

DETAILED DESCRIPTION

This description is not to be taken in a limiting sense, but is mademerely for the purpose of illustrating the general principles of theinvention. The section titles and overall organization of the presentdetailed description are for the purpose of convenience only and are notintended to limit the present invention.

Definitions

As used herein the specification, “a” or “an” may mean one or more. Asused herein in the claim(s), when used in conjunction with the word“comprising”, the words “a” or “an” may mean one or more than one. Asused herein “another” may mean at least a second or more.

The term “preventing” is art-recognized, and when used in relation to acondition, such as a neurodegenerative disease, is well understood inthe art, and includes administration of a composition which reduces thefrequency or severity of the symptoms of a medical condition in asubject relative to a subject who does not receive the composition.Thus, the prevention of neurodegenerative disease progression includes,for example, reducing the average amount of gray matter loss in apopulation of patients receiving an estrogen receptor beta ligandrelative to a control population that did not receive the estrogenreceptor beta ligand e.g., by a statistically and/or clinicallysignificant amount. Similarly, the prevention of neurodegenerativedisease progression includes reducing the likelihood that a patientreceiving an estrogen receptor beta ligand will develop as much graymatter loss as the patient would develop if the patient did not receivethe estrogen receptor beta ligand. Similarly, the prevention ofneurodegenerative disease progression includes reducing the likelihoodthat a patient receiving an estrogen receptor beta ligand will develop adisability, such as cognitive decline, learning disability, and/ormemory loss, as much as the patient would develop if the patient did notreceive the estrogen receptor beta ligand.

The term “subject” as used herein refers to a living mammal and may beinterchangeably used with the term “patient”. In certain embodiments,the subject is a rodent, such as a mouse, or a primate, such as a human.The subject may be a male or female. In some embodiments, the subject isa female.

The terms “substantial brain gray matter atrophy” and “substantial lossof brain gray matter volume” as used herein, refers to a patient havinggreater than about 0.5% brain gray matter loss per annum (e.g., about0.5%, or about 0.6%, or about 0.7%, or about 0.8%, or about 0.9%, orabout 1.0%, or about 1.1%, or about 1.2%, or about 1.3%, or about 1.4%,or about 1.5%, or about 2.0% brain gray matter loss per annum). Forexample “substantial brain gray matter atrophy” and “substantial loss ofbrain gray matter volume” may refer to a patient having a loss of braingray matter of greater than about 0.3% in a period of at least about 6months (e.g., about 0.4%, or about 0.5%, or about 0.6%, or about 0.7%,or about 0.8%, or about 0.9%, or about 1.0%, or about 1.1%, or about1.2%, or about 1.3%, or about 1.4%, or about 1.5%, or about 2.0 in aperiod of about 6 months, or about 12 months, or about 18 months, orabout 24 months). The amount of gray matter loss may be determined, forexample, by using an imaging technique or surrogate marker.

As used herein, the term “treating” or “treatment” includes reducing,arresting, or reversing the symptoms, clinical signs, or underlyingpathology of a condition in a manner to stabilize or improve a subject'scondition or to prevent the likelihood that the subject's condition willworsened as much as if the subject did not receive the treatment.

Generally, the invention involves a method of treating a mammalexhibiting neurodegeneration, such as clinical symptoms of an autoimmuneor neurodegenerative disease, comprising administering a primary agentbeing an estrogen receptor ligand. In one embodiment, the estrogenreceptor ligand is an ERβ ligand. Optionally, a secondary agent may beused to treat neurodegeneration, preferably in a lower dose than if notused in combination with an estrogen receptor ligand. The treatment isaimed at providing a protective effect after the acute phase, reducingthe number and/or degree of signs and symptomology and/or progression ofa neurodegenerative disease.

The beneficial effect of treatment can be evidenced by a protectiveeffect on the progression of disease symptomology after the acute phase,a reduction in the number and/or severity of some or all of the clinicalsigns and/or symptoms, or an improvement in the overall health.

For example, patients who have clinical symptoms of a neurodegenerativedisease often suffer from some or all of the following symptoms:worsening of pre-existing symptoms (memory loss in Alzheimer's disease),the appearance of new symptoms (e.g. balance problems in Parkinson'sdisease) or increased generalized weakness and fatigue. MS patients inparticular suffer from the following symptoms: weakness, numbness,tingling, loss of vision, memory difficulty, learning disability, andextreme fatigue. Thus, an amelioration of disease in MS would include areduction in the frequency or severity of onset of weakness, numbness,tingling, loss of vision, memory difficulty, learning disability, andextreme fatigue. On imaging of the brain (MRI) amelioration or reducedprogression of disease would be evidenced by a slowing in the rate ofcerebral, cerebellar or spinal cord atrophy formation or a stabilizationor slowing of the accumulation of T2 lesions.

Patients may also express criteria indicating they are at risk fordeveloping neurodegenerative diseases. These patients may bepreventatively treated to delay the onset of clinical signs and/orsymptomology. More specifically, patients who present initially withclinically isolated syndromes (CIS) may be treated using the treatmentparadign outlined in this invention. These patients have had at leastone clinical event consistent with MS, but have not met full criteriafor MS diagnosis since the definite diagnosis requires more than oneclinical event at another time. Treatment of the present invention wouldbe advantageous at least in providing a protective effect after theacute phase of clinically definite MS.

PRIMARY AGENT. The primary agents useful in this invention are estrogenreceptor β agonists. These agonists may be steroidal or non-steroidalagents which bind to and/or cause a change in activity or binding of theestrogen receptor β. In certain preferred embodiments, the estrogenreceptor β agonist agonizes estrogen receptor β with an EC₅₀ at least 10times lower than its EC₅₀ for agonizing estrogen receptor α, preferablyat least 100 times lower.

One agent useful in this invention alone or in combination is an ERβligand, known as AC-186 or compounds substantially similar in structureand function thereto (see the compounds disclosed in PCI PatentPublication WO 2013/017619 A1, incorporated herein by reference), asevidenced at least by treating in the EAE models described below.

SECONDARY ACTIVE AGENTS. Any one or a combination of secondary activeagents may be included in combination with the primary agent.Alternatively, any one or a combination of secondary active agents maybe administered independently of the primary agent, but concurrent intime for exposure to at least two agents for the treatment of theneurodegenerative disease. Alternatively, the primary and secondaryagents may be used in an alternating fashion to prevent or treatneurodegeneration.

The secondary agents are preferably immunotherapeutic agents, which actsynergistically with the primary agent to diminish the symptomology ofthe neurodegenerative disease. Secondary active agents may be selectedto enhance the effect of the primary agent, or affect a different systemthan that affected by the primary agent.

The secondary agent may be selected from the group comprisingβ-interferon compounds. Examples include as β-interferon (e.g., Avonex®(interferon-beta 1a), Rebif® (by Scrono); Extavia®, Betaseron®(interferon-beta 1b; Berlex, Schering), PLEGRIDY™ (peginterferonbeta-1a; Biogen)). Optionally, the following agents may be used:dimethyl fumarate (Tecfidera®; BG-12), fingolimod (Gilenya®), glatirameracetate (Copaxone®), interferon beta-1a (e.g., Avonex® or Rebif®),pegylated interferon-beta-1a (PLEGRIDY™; Biogen) mitoxantrone(Novantrone®; Lederle Labs), natalizumab (Tysabri®), anti-LINGO-1antibody (BIIB033, Biogen-Idec), Antegren® (Elan Corp.), teriflunomide(Aubagio®), mycophenolate mofetil (CellCept® Hoffman-LaRoche Inc.),paclitaxel (Taxol®; Bristol-Meyers Oncology), cyclosporine (such ascyclosporin A), corticosteroids (e.g., prednisone, methylprednisolone),azathioprine, cyclophosphamide, methotrexate, cladribine,4-aminopyridine, and tizanidine.

Surprisingly, the estrogen receptor beta ligand of formula I (AC-186)reduces macrophage activity in the brain and spinal cord: thus, theestrogen receptor beta ligand of formula I and other compounds ofsimilar activity may be used as an immunotherapeutic oranti-inflammatory agent. Accordingly, the dose of a secondary agent withimmunotherapeutic or anti-inflammatory activity may be reduced whenadministered in combination with the estrogen receptor beta ligand offormula I, for example, to reduce the occurrence or severity of unwantedside effects associated with the secondary agent. Similarly, theadditional agent may be administered at a lower dose due to thesynergistic effect with the combination of the first and second agents.Examples include a glucocorticoid, precursor, analog or glucocorticoidreceptor agonist or antagonist. For example, prednisone may beadministered, most preferably in the dosage range of about 5-60milligrams per day. Also, methyl prednisone (Solu-Medrol) may beadministered, most preferably in the dosage range of about 1-2milligrams per day. Glucocorticoids are currently used to treat relapseepisodes in MS patients within this dosage range.

The dose of the disease-modifying therapeutic may be decreased when usedin combination with the estrogen receptor beta ligand. For example, acurrent standard dose for glatiramer acetate (Copaxone®) is 40 mgsubcutaneously (s.c.) three times a week, or 20 mg s.c. daily. Inconjunction with the estrogen receptor beta ligand in accordance withthe invention, the dose for glatiramer acetate (Copaxone®) may bereduced by up to 10 percent, by up to 15 percent, by up to 20 percent,by up to 25 percent, by up to 30 percent, by up to 35 percent, by up to40 percent, by up to 45 percent, by up to 50 percent or more, e.g., to20 mg s.c. six times a week, five times a week, four times a week, orthree times a week.

As another example, a current standard dose for fingolimod (Gilenya®) is0.5 mg by mouth (p.o.) daily. In conjunction with the estrogen receptorbeta ligand in accordance with the invention, the dose for fingolimod(Gilenya®) may be reduced by up to 10 percent, by up to 15 percent, byup to 20 percent, by up to 25 percent, by up to 30 percent, by up to 35percent, by up to 40 percent, by up to 45 percent, by up to 50 percentor more. e.g., to 0.45, 0.40, 0.35, 0.30, or 0.25 mg p.o. daily.

As another example, a current standard dose for dimethyl fumarate(Tecfidera®) is 240 mg p.o. daily. In conjunction with the estrogenreceptor beta ligand in accordance with the invention, the dose fordimethyl fumarate (Tecfidera®) may be reduced by up to 10 percent, by upto 15 percent, by up to 20 percent, by up to 25 percent, by up to 30percent, by up to 35 percent, by up to 40 percent, by up to 45 percent,by up to 50 percent or more, e.g., to 220 mg p.o. daily, 200 mg p.o.daily, 180 mg p.o. daily, 160 mg p.o. daily, 150 mg p.o. daily, 140 mgp.o. daily, or 120 mg p.o. daily.

As yet another example, a current standard dose for interferon beta-1a(Avonex® or Rebif®) is 30 μg intramuscularly (i.m.) weekly (Avonex®) or44 μg s.c. three days a week (Rebif®). In conjunction with the estrogenreceptor beta ligand in accordance with the invention, the dose forAvonex® may be reduced to 27, 26, 25, 24, 23, 22, 21, 20, 19, 18, 17,16, or 15 μg i.m. weekly, or 30 μg intramuscularly (i.m.) every 8, 9, 10or 11 days, and the dose for Rebif® may be reduced to 40, 39, 38, 37,36, 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, 24, 23, or 22 μg s.c.three days a week, or 44 μg s.c. two days a week.

As yet another example, a current standard dose for interferon beta-1b(Betaseron® or Extavia®) is 0.25 mg s.c. every other day (Betaseron® orExtavia®). In conjunction with the estrogen receptor beta ligand inaccordance with the invention, the dose for interferon beta-1b(Betaseron® or Extavia®) may be reduced to 0.225, 0.200, 0.180, 0.175,0.170, 0.160, 0.150, 0.140, 0.130, or 0.125 mg s.c. every other day, or0.25 mg s.c. every third day.

THERAPEUTICALLY EFFECTIVE DOSAGE OF THE PRIMARY AGENT. A therapeuticallyeffective dose of the primary agent is one sufficient to raise the serumconcentration above basal levels, and preferably to produce a biologicaleffect in the central nervous system, with no off target binding effectson breast or uterus.

In certain embodiments where the primary agent is AC-186, the dose maybe about 1-100 mg/kg/day, but can be titrated by one of skill in the artto achieve the desired benefits while reducing the risks including sideeffects of this therapy. In some embodiments, the primary agent isAC-186, and the compound is administered at a dose sufficient to achievea mean blood concentration of the compound between 1 ng/ml and 1000ng/ml. In other embodiments, the compound is administered at a dosesufficient to achieve a mean blood concentration of the compound between100 ng/ml and 200 ng/ml.

The dosage of the primary agent may be selected for an individualpatient depending upon the route of administration, severity of disease,age and weight of the patient, other medications the patient is takingand other factors normally considered by the attending physician, whendetermining the individual regimen and dosage level as the mostappropriate for a particular patient.

The use of this group of primary agents is advantageous in at least thatother known or experimental treatments for MS are chemotherapeuticimmunosuppressants which have significant risks and side effects topatients, including decreasing the ability of the patient to fightinfections, inducing liver or heart toxicity which are not caused byestrogen treatment. Other agents used in MS do not cause these sideeffects, but are associated with flu-like symptoms or chest tightness.Further, these previously used agents are associated with local skinreactions since they entail injections at frequencies ranging from dailyto once per week.

DOSAGE FORM. The therapeutically effective dose of the primary agentincluded in the dosage form is selected at least by considering theprimary agent selected and the mode of administration, preferably oral.The dosage form may include the active primary agent in combination withother inert ingredients, including adjutants and pharmaceuticallyacceptable carriers for the facilitation of dosage to the patient asknown to those skilled in the pharmaceutical arts. The dosage form maybe any form suitable to cause the primary agent to enter into thetissues of the patient.

In certain embodiments, the primary agent is formulated for buccal orsublingual administration. For example, the dosage form of the primaryagent may be a tablet, drop, spray, film, thin-film, or lozenge.

In one embodiment, the dosage form of the primary agent is an oralpreparation (liquid, tablet, capsule, caplet or the like) which whenconsumed results in elevated levels of the primary agent in blood serum.The oral preparation may comprise conventional carriers includingdiluents, binders, time release agents, lubricants and disintegrants.

Possible oral administration forms are all the forms known from theprior art such as, tablets, dragees, pills or capsules, which areproduced using conventional adjuvants and carrier substances. In thecase of oral administration it has provided appropriate to place thedaily units, which in case comprise a combination of the primary andsecondary agents, in a spatially separated and individually removablemanner in a packaging unit, so that it is easy to check whether thetypically daily taken, oral administration form has in fact been takenas it is important to ensure that there are no taking-free days.

In other embodiments of the invention, the dosage form may be providedin a topical preparation (lotion, crèmc, ointment, patch or the like)for transdermal application. Alternatively, the dosage form may beprovided in a suppository or the like for intravaginal or transrectalapplication. Alternatively, the agents may be provided in a form forinjection or for implantation.

In the transdermal administration of the combination according to theinvention, the agents may be applied to a plaster or also can be appliedby transdermal, therapeutic systems and are consequently supplied to theorganism. For example, an already prepared combination of the agents orthe latter individually can be introduced into such a system, which isbased on ionotherapy or diffusion or optionally a combination of theseeffects.

That the agents can be delivered via these dosage forms is advantageousin that currently available therapies, for MS for example, many areinjectables, which are inconvenient for the user and lead to decreasedpatient compliance with the treatment. Non-injectable dosage forms arefurther advantageous over current injectable treatments which oftencause side effects in patients including flu-like symptoms(particularly, P interferon) and injection site reactions which may leadto lipotrophy (particularly, glatiramer acetate copolymer-1). Currentlyavailable oral treatments for MS, however, consist of onlyanti-inflammatory agents, which display only a modest effect ondisability and are only effective in the early stages of the disease.

However, in additional embodiment, the dosage form may also allow forpreparations to be applied subcutaneously, intravenously,intramuscularly or via the respiratory system.

DOSE. By way of example, which is consistent with the currenttherapeutic uses for these treatments. Avonex00 in a dosage of about 0to about 30 meg may be injected intramuscularly once a week. Betascron®in a dosage of about 0 to about 0.25 mg may be injected subcutaneouslyevery other day. Copaxone® in a dosage of about 0 to about 20 mg may beinjected subcutaneously every day. Finally, Rebif® may be injected at atherapeutic dose and at an interval to be determined based on clinicaltrial data. One objective would be to select the minimal effective doseof β-interferon given the side effects, injection site reactions andcompliance issues associated with its use. Thus, the second agent may beadministered at a reduced dose or with reduced frequency due tosynergistic effects with the primary agent. However, dosages and methodof administration may be altered to maximize the effect of thesetherapies in conjunction with estrogen β receptor ligand treatment.Dosages may be altered using criteria that are known to those skilled inthe art of diagnosing and treating autoimmune diseases.

In certain aspects, the invention relates to methods of slowing orhalting gray matter atrophy in a neurodegenerative disease patient,comprising administering to the patient an estrogen receptor betaligand.

In certain embodiments, the invention relates to methods of reversinggray matter atrophy in a neurodegenerative disease patient, comprisingadministering to the patient an estrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of slowing orhalting progression of disability in a neurodegenerative diseasepatient, comprising administering to the patient an estrogen receptorbeta ligand.

In certain embodiments, the invention relates to methods of reversingprogression of disability in a neurodegenerative disease patient,comprising administering to the patient an estrogen receptor betaligand.

In certain embodiments, the invention relates to methods of preventingthe progression of a neurodegenerative disease, comprising administeringto a patient presenting with cognitive disability an effective amount ofan estrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treatingneurodegenerative disease in a patient having cognitive disability,comprising administering to the patient an effective amount of anestrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treatingand/or slowing the progression of a neurodegenerative disease,comprising evaluating a patient's cognitive ability, and administeringan effective amount of an estrogen receptor beta ligand to a patientsuffering from cognitive disability.

In certain embodiments, the invention relates to methods of slowing orhalting the progression of memory loss in a neurodegenerative diseasepatient, comprising administering to the patient an estrogen receptorbeta ligand.

In certain embodiments, the invention relates to methods of reversingthe progression of memory loss in a neurodegenerative disease patient,comprising administering to the patient an estrogen receptor betaligand.

In certain embodiments, the invention relates to methods of preventingthe progression of a neurodegenerative disease, comprising administeringto a patient presenting with memory loss an effective amount of anestrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treatingneurodegenerative disease in a patient having substantial memory loss,comprising administering to the patient an effective amount of anestrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treatingand/or slowing the progression of a neurodegenerative disease,comprising evaluating a patient's memory, and administering an effectiveamount of an estrogen receptor beta ligand to a patient suffering frommemory loss.

In certain embodiments, the invention relates to methods of slowing orhalting the progression of a learning disability in a neurodegenerativedisease patient, comprising administering to the patient an estrogenreceptor beta ligand.

In certain embodiments, the invention relates to methods of reversingthe progression of a learning disability in a neurogenerative diseasepatient, comprising administering to the patient an estrogen receptorbeta ligand.

In certain embodiments, the invention relates to methods of preventingthe progression of a neurodegenerative disease, comprising administeringto a patient presenting with a learning disability an effective amountof an estrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treatingneurodegenerative disease in a patient having a learning disability,comprising administering to the patient an effective amount of anestrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treatingand/or slowing the progression of a neurodegenerative disease,comprising evaluating a patient's learning ability, and administering aneffective amount of an estrogen receptor beta ligand to a patientsuffering from learning disability.

In certain embodiments, the invention relates to methods of preventingthe progression of a neurodegenerative disease, comprising administeringto a patient presenting with brain gray matter atrophy an effectiveamount of an estrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treatingneurodegenerative disease in a patient having substantial brain graymatter atrophy, comprising administering to the patient an effectiveamount of an estrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treatingand/or slowing the progression of a neurodegenerative disease,comprising evaluating a patient's brain gray matter, and administeringan effective amount of an estrogen receptor beta ligand to a patientsuffering from brain gray matter atrophy.

In certain embodiments, the invention relates to methods of treating orpreventing a neurodegenerative disease in a patient having greater thanabout 0.1% brain gray matter loss per annum, comprising administering tothe patient an effective amount of an estrogen receptor beta ligand. Incertain embodiments, the invention relates to methods of treating orpreventing a neurodegenerative disease in a patient having greater thanabout 0.5% brain gray matter loss per annum, comprising administering tothe patient an effective amount of an estrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treating aneurodegenerative disease in a patient having a loss of brain graymatter of greater than about 0.3% in a period of at least about 6months, comprising administering to the patient an effective amount ofan estrogen receptor beta ligand.

In certain embodiments, the invention relates to methods of treating aneurodegenerative disease in a patient who is non-responsive totreatment with a multiple sclerosis therapy, comprising administering tothe patient an effective amount of an estrogen receptor beta ligand,wherein the patient is classified as non-responsive based on asubstantial loss of brain gray matter volume during treatment with thefirst multiple sclerosis treatment agent.

In some embodiments, the brain gray matter loss per annum is about 0.2%,or about 0.3%, or about 0.4%, or about 0.5%, or about 0.6%, or about0.7%, or about 0.8%, or about 0.9%, or about 1.0%, or about 1.1%, orabout 1.2%, or about 1.3%, or about 1.4%, or about 1.5%, or about 2.0%.In some embodiments, the loss of brain gray matter is about 0.4%, orabout 0.5%, or about 0.6%, or about 0.7%, or about 0.8%, or about 0.9%,or about 1.0%, or about 1.1%, or about 1.2%, or about 1.3%, or about1.4%, or about 1.5%, or about 2.0 in a period of about 6 moths, or about12 months, or about 18 months, or about 24 months. The gray matter lossmay be located in one or more of total brain, cerebral cortex,cerebellum, thalamus, caudate nucleus, and putamen.

The gray matter loss may be measured using an imaging technique orsurrogate marker. In some embodiments, the gray matter loss is measuredusing an imaging technique selected from magnetic resonance imaging(MRI), fast fluid-attenuated inversion recovery (FLAIR), doubleinversion recovery (DIR), phase-sensitive inversion recovery (PSIR),ultra high-field MRI, magnetization transfer imaging (MTI),T-relaxometry, diffusion tensor imaging (DTI), proton magnetic resonancespectroscopy (MRS)), and related techniques and combinations thereof.The gray matter loss may be measured using a surrogate marker selectedfrom one or more of nogo receptor, kallikrein-6 (neurosin),cerebellin-1, ceruloplasmin, dickkopf-3 (rig-like 7-1), amyloid betaprecursor-like protein 1, activated leukocyte cell adhesion molecule(CD166), neural cell adhesion molecule 2, neural epidermal growth factorlike 2/cerebral protein-12, clusterin (apolipoprotein j, complementlysis inhibitor), brevican, neuronal cadherin, chitinase-3-like 1protein, neogenin, multifunctional protein MFP (collagen alpha-1 (XVIII)chain; endostatin), dystroglycan 1, contactin 2, ephrin type a receptor4, neural cell adhesion molecule L1 like protein, and contactin 1.

In some embodiments, the treatment comprises one or more of preventingdisease progression, slowing of disease progression, reducing the numberof disease relapses or clinical exacerbations, and slowing theaccumulation of physical disability.

In some embodiments, the patient has one or more of: experienced a firstclinical episode, MRI features consistent with multiple sclerosis, aninadequate response to an alternate MS therapy, and an inability totolerate an alternate MS therapy.

In certain embodiment, the method is a method for slowing, halting, orreversing progression of a cognitive or physical disability in a subjectwith a neurodegenerative disease, comprising identifying a subject whohas experienced progression of a cognitive or physical disability andinitiating treatment of the subject by a method as described herein. Incertain embodiments, the method further comprises testing the severityof the subject's cognitive or physical disability to determine a scorerepresentative of the state of the subject's cognitive or physicaldisability after receiving the treatment for at least about six months,and, optionally, comparing the score to a score determined for thesubject prior to or at about the time of initiating the treatment.

In certain embodiments, the method is a method for slowing, halting, orreversing the progression of memory loss in a subject with aneurodegenerative disease, comprising identifying a subject who hasexperienced progression of memory loss and initiating treatment of thesubject by a method as described herein. In certain embodiments, themethod further comprises testing the severity of the subject's memoryloss to determine a score representative of the state of the subject'smemory after receiving the treatment for at least about six months, and,optionally, comparing the score to a score determined for the subjectprior to or at about the time of initiating the treatment.

In certain embodiments, the method is a method for slowing, halting, orreversing the progression of a learning disability in a subject with aneurodegenerative disease, comprising identifying a subject who hasexperienced progression of a learning disability and initiatingtreatment of the subject by a method as described herein. In certainembodiments, the method further comprises testing the severity of thesubject's learning disability to determine a score representative of thestate of the subject's learning ability after receiving the treatmentfor at least about six months, and, optionally, comparing the score to ascore determined for the subject prior to or at about the time ofinitiating the treatment.

The various methods disclosed herein can be methods for improvingwalking, vision, balance, cognition, learning, memory, or other symptomsin a subject, such as a subject with multiple sclerosis, and/or methodsfor improving multiple sclerosis functional composite (MSFC). EDSS, 7/24test, or MSSS scores in a subject, such as a subject with multiplesclerosis. Thus, in certain embodiments, the methods of treatmentdisclosed herein include methods for stabilizing or improving disabilityin a patient, whereby the patient's disability score (as measured byeither of these tests or another suitable test) after six months, oneyear, or two years of therapy is at least about 10%, at least about 25%,at least about 40%, at least about 50%, or even at least about 60%higher relative to a control patient not receiving the estrogen receptorbeta ligand (but otherwise receiving the same treatment as the patient).Alternatively, the patient's disability score (as measured by either ofthese tests or another suitable test) after six months, one year, or twoyears of therapy is within about 2% or within about 5% of an earlierassessment, or at least about 2%, at least about 5%, at least about atleast about 10%, at least about 25%, at least about 40%, at least about50%, or even at least about 60% higher than the earlier assessment.

For example, progression of a walking disability can be tested using awalking test, e.g., assessing the subject's performance on a 25-footwalk test at different points in time, such as at 0 months (baseline), 6months, 1 year, and 2 years. In certain embodiments, if there isdocumented worsening in walking (takes more seconds) by 20 percent ascompared to baseline (optionally if this worsening is confirmed on asubsequent walk test (e.g., 3 months later)), then the subject is deemedto have progressive worsening in walking. For such a patient not alreadyreceiving an estrogen receptor beta ligand therapy, the subjectdemonstrating the progressive walking disability commences treatmentwith an estrogen receptor beta ligand. The walking test may be repeated(e.g., at 1 year and/or 2 years from the start of an estrogen receptorbeta ligand treatment) to assess whether the treatment slowed or haltedany further worsening in walking performance, e.g., as measured by thewalking test.

Improvements in cognition outcomes associated with MS therapy, whetherslowing of cognitive decline, stabilization of cognitive decline, orimprovement of cognitive function, can be assessed using the PASAT(e.g., PASAT 2 or PASAT 3) or SDMT test, or alternatively the MS-COGtest (see Erlanger et al., J Neuro Sci 340: 123-129 (2014)). Thus, incertain embodiments, the methods of treatment disclosed herein includemethods for stabilizing or improving cognition in a patient, whereby thepatient's cognition outcome after one year of therapy is at least about2%, at least about 5%, at least about 10%, at least about 25% at leastabout 40%, at least about 50%, or even at least about 60% higherrelative to a control patient not receiving the estrogen receptor betaligand therapy (but otherwise receiving the same treatment as theestrogen receptor beta ligand patient), e.g., as measured by any of thepreceding tests. Alternatively, the patient's cognition outcome aftersix months, one year, or two years of therapy may be within about 2% orwithin about 5% of an earlier assessment, or at least about 2%, at leastabout 5%, at least about 10%, at least about 25%, at least about 40%, atleast about 50%, or even at least about 60% higher than the earlierassessment, e.g., as measured by any of the preceding tests at differenttimes.

Improvements in memory associated with MS therapy, whether slowing ofmemory loss, stabilization of memory loss, or improvement of memory, canbe assessed using, for example, the 7/24 Spatial Recall Test. Thus, incertain embodiments, the method % of treatment disclosed herein includemethods for stabilizing memory loss or improving memory in a patient,whereby the patient's memory after one year of therapy is at least about10%, at least about 25%, at least about 30%, at least about 40%, atleast about 45%, at least about 50%, or even at least about 60% higherrelative to a control patient not receiving the estrogen/progestogentherapy (but otherwise receiving the same treatment as theestrogen-treated patient), e.g., as measured by the 7/24 Spatial RecallTest.

In some embodiments, substantial loss of memory refers to less thanperfect performance or worsening performance on the 7/24 Spatial RecallTest or Delayed Recall Test. For example, a subject who scores less than7 on the Spatial Recall Test or Delayed Recall Test has substantialmemory loss. Additionally, a subject who scores a 7 on the 7/24 SpatialRecall Test or Delayed Recall Test on a first date followed by a scoreof 6 or less on a subsequent date has substantial loss of memory overthe period of time defined by the first date and the subsequent date.

For example, a subject who scores below 50 on PASAT (and optionally ifsuch low score is verified upon a second subsequent test, such as withinone week to one month of the first) may be deemed to have cognitivedisability. For such a patient not already receiving treatment, thesubject demonstrating the cognitive disability may commence treatmentwith an estrogen receptor beta ligand. In certain embodiments, thecognitive test may be repeated (e.g., at about six months from the startof treatment) to assess whether the treatment slowed or halted anyfurther worsening in cognitive performance, e.g., as measured by thePASAT test. In certain such embodiments, the patient's score mayincrease by at least 3 points over the course of six to twelve months ofthe estrogen receptor beta ligand therapy.

The estrogen receptor beta ligand may be a compound having the structureof formula I:

The compound of formula I is called AC-186. In some embodiments, theestrogen receptor beta ligand is a compound that is substantiallysimilar in structure and function to AC-186, such as one of thecompounds disclosed in U.S. Patent Application Publication Nos.2009/0131510 and 2014/0275284, and PCT Patent Application PublicationNos. WO 2013/017619 and WO 2014/125121 (hereby incorporated byreference, especially for the molecules disclosed therein).

In some embodiments, the ER ligand is a compound selected from thecompounds disclosed in U.S. Patent Application Publication Nos.2012/0128435, 2012/0202853, 2012/0202861, 2013/0131061, or 2014/0323518and PCT Patent Application Publication Nos. WO 2012/022776 and2012/136772 (hereby incorporated by reference, especially for themolecules disclosed therein). In some embodiments, the ERβ ligand isKBRV1 or KBRV2 (Karo Bio, Huddinge, Sweden).

The ERβ ligand may be any one of the following compounds:

In some embodiments, the ERβ ligand is Compound II, III, or IV.

The ERβ ligand may be5-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1-(4-hydroxyphenyl)-3-methyl-1H-pyrazole-4-carboximidamide;5-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-3-methyl-1H-pyrazole-4-carboxamide;5-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-3-methyl-1H-pyrazole-4-carbaldehydeoxime;5-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-3-propyl-1H-pyrazole-4-carbaldehydeoxime;5-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1-(4-hydroxyphenyl)-3-propyl-1H-pyrazole-4-carboximidamide;5-(2,5-dimethyl-1lH-pyrrol-1-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;5-(diethylamino)-3-ethyl-N′-hydroxy-1-(4-hydroxyphenyl)-1H-pyrazole-4-carboximidamide;5-(1,3-dimethyl-1H-pyrrol-2-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;5-(5-bromo-1,3-dimethyl-1H-pyrrol-2-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;3-ethyl-N′-hydroxy-1-(4-hydroxyphenyl)-5-(3-methylfuran-2-yl)-1H-pyrazole-4-carboximidamide;4-(4-(3,5-dimethylisoxazol-4-yl)-5-(hydroxymethyl)-1-methyl-1H-pyrazol-3-yl)phenol;4-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carbaldehydeoxime;4-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carboximidamide;4-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carboxamide;4-(4-(3,5-dimethylisoxazol-4-yl)-5-(2-hydroxyethyl)-1-methyl-1H-pyrazol-3-yl)phenol;4-(2,6-difluorophenyl)-N′-hydroxy-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carboximidamide;5-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-3-methyl-1H-pyrazole-4-carbonitrile;5-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-3-isopropyl-1H-pyrazole-4-carbaldehydeoxime;5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-1-(4-hydroxyphenyl)-1H-pyrazole-4-carbaldehydeoxime;5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-N′-hydroxy-1-(4-hydroxyphenyl)-1H-pyrazole-4-carboximidamide,1-(3,5-difluoro-4-hydroxyphenyl)-5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-N′-hydroxy-1H-pyrazole-4-carboximidamide;5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-1-(3-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-pyrazole-4-carboximidamide;1-(2,3-difluoro-4-hydroxyphenyl)-5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-N′-hydroxy-1H-pyrazole-4-carboximidamide;5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-pyrazole-4-carboximidamide;1-(2,3-difluoro-4-hydroxyphenyl)-5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-1H-pyrazole-4-carbaldehydeoxime (isomer A);1-(2,3-difluoro-4-hydroxyphenyl)-5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-1H-pyrazole-4-carbaldehydeoxime (isomer B);5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-1-(2-fluoro-4-hydroxyphenyl)-1H-pyrazole-4-carbaldehydeoxime;1-(3,5-difluoro-4-hydroxyphenyl)-5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-H4-pyrazole-4-carbaldehydeoxime;1-(2,5-difluoro-4-hydroxyphenyl)-5-(3,5-dimetiylisoxazol-4-yl)-3-ethyl-N′-hydroxy-1H-pyrazole-4-carboximidamide;1-(2,5-difluoro-4-hydroxyphenyl)-5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-1H-pyrazole-4-carbaldehydeoxime;5-(3,5-dimethylisoxazol-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;5-(3,5-dimethylisoxazol-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-propyl-1H-pyrazole-4-carboximidamide;5-(3,5-dimethylisoxazol-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-3-methyl-1H-pyrazole-4-carbaldehydeoxime;1-(2,3-difluoro-4-hydroxyphenyl)-3-ethyl-5-(1-methyl-1H-pyrrol-2-yl)-1H-pyrazole-4-carbaldehydeoxime;2-(2-fluoro-4-hydroxyphenyl)-2′,4′,5-trimethyl-2H,2′H-3,3′-bipyrazole-4-carbaldehydeoxime;2-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2′,4′,5-trimethyl-2H,2′H-3,3′-bipyrazole-4-carboximidamide;5-(2,6-dimethylphenyl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;5-(2,5-dimethyl-1H-pyrrol-1-yl)-3-ethyl-N′-hydroxy-1-(4-hydroxyphenyl)-1H-pyrazole-4-carboximidamide;5-(2,6-dimethylphenyl)-1-(2-fluoro-4-hydroxyphenyl)-3-methyl-1H-pyrazole-4-carbaldehydeoxmie;5-(2,5-dimethyl-1H-pyrrol-1-yl)-1-(3-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide:1-(2,3-difluoro-4-hydroxyphenyl)-5-(2,5-dimethyl-1H-pyrrol-1-yl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;1-(3,5-difluoro-4-hydroxyphenyl)-5-(2,5-dimethyl-1H-pyrrol-1-yl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;3-(3-chloro-4-hydroxyphenyl)-4-(3,5-dimethylisoxazol-4-yl)-1-methyl-1H-pyrazole-5-carboxamide;4-(2,6-difluorophenyl)-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carbaldehydeoxime;4-(2,6-dichlorophenyl)-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carbaldehydeoxime;4-(2,6-dichlorophenyl)-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carboxamide;4-(2,6-dichlorophenyl)-N′-hydroxy-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carboximidamide;4-(3,5-dimethylisoxazol-4-yl)-3-(2-fluoro-4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carbaldehydeoxime;4-(3,5-dimethylisoxazol-4-yl)-3-(2-fluoro-4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carboxamide;4-(3,5-dimethylisoxazol-4-yl)-3-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1-methyl-1H-pyrazole-5-carboximidamide;5-((Z)-but-2-en-2-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;5-(2,4-dimethylthiophen-3-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;5-(3,5-dimethylpyridin-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-3-methyl-1H-pyrazole-4-carboximidamide;5-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1-(4-hydroxy-2-methylphenyl)-3-methyl-1H-pyrazole-4-carboximidamide;5-(3,5-dimethylisoxazol-4-yl)-3-ethyl-N′-hydroxy-1-(4-hydroxy-2-methylphenyl)-1H-pyrazole-4-carboximidamide;5-(2,5-dimethyl-1H-pyrrol-1-yl)-N′-hydroxy-1-(4-hydroxy-2-methylphenyl)-3-methyl-1H-pyrazole-4-carboximidamide;N′-hydroxy-1-(4-hydroxy-2-methylphenyl)-3-methyl-5-(2-methyl-5-propyl-1H-pyrrol-t-yl)-1ii-pyrazole-4-carboximidamide;4-(2,4-dimethylthiophen-3-yl)-3-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1-methyl-1H-pyrazole-5-carboximidamide;4-(3,5-dimethylisoxazol-4-yl)-3-(3-fluoro-4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carbaldehydeoxime;4-(3,5-dimethylisoxazol-4-yl)-3-(3-fluoro-4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carboxamide:4-(3,5-dimethylisoxazol-4-yl)-3-(3-fluoro-4-hydroxyphenyl)-N′-hydroxy-1-methyl-1H-pyrazole-5-carboximidamide;(Z)-4-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxy-2-methylphenyl)-1-methyl-1H-pyrazole-5-carbaldehydeoxime;(E)-4-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxy-2-methylphenyl)-1-methyl-1H-pyrazole-5-carbaldehydeoxime;4-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-3-(4-hydroxy-2-methylphenyl-1-methyl-1H-pyrazole-5-carboximidamide;4-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxy-2-methylphenyl)-1-methyl-1H-pyrazole-5-carboxamide;4-(2,4-dimethylthiophen-3-yl)-N-hydroxy-3-(4-hydroxy-2-methylphenyl)-1-methyl-1H-pyrazole-5-carboximidamide;4-(3,5-dimethylisothiazol-4-yl)-3-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1-methyl-1H-pyrazole-5-carboximidamide;4-(2,6-dimethylphenyl)-3-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1-methyl-1H-pyrazole-5-carboximidamide;4-(2,4-dimethylfuran-3-yl)-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carbaldehydeoxime;4-(3,5-dimethylisothiazol-4-yl)-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carbaldehydeoxime; or4-(3,5-dimethylisothiazol-4-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1-methyl-1H-pyrazole-5-carboximidamide.

The ERβ ligand may be2-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxyphenyl)-1H-indole-1-carbonitrile:2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1H-indole-1-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxyphenyl)-1H-indole-1-carboxamide;2-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxyphenyl)-N,N-dimethyl-1H-indole-1-sulfonamide;2-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxyphenyl)-1H-indole-1-carbaldehydeoxime;4-(2-(3,5-dimethylisoxazol-4-yl)-1-(methylsulfonyl)-1H-indol-3-yl)phenol;2-((Z)-but-2-en-2-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1H-indole-1-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-N-ethyl-3-(4-hydroxyphenyl)-1H-indole-1-carboxamide;2-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxyphenyl)-N-methyl-1H-indole-1-carboxamide;2-(2,4-dimethylthiophen-3-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1H-indole-1-carboximidamide;2-(2,4-dimethylthiophen-3-yl)-3-(4-hydroxyphenyl)-1H-indole-1H-carboxamide;2-(2,6-dimethylphenyl)-N′-hydroxy-3-(4-hydroxyphenyl-1H-indole-1H-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxyphenyl)-N-isopropyl-1H-indole-1-carboxamide;2-(3,5-dimethylisoxazol-4-yl)-3-(4-hydroxyphenyl)-N-pentyl-1-indole-1-carboxamide;2-(2,4-dimethylthiophen-3-yl)-N-ethyl-3-(4-hydroxyphenyl)-1H-indole-1-carboxamide:3-(3,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1H-indole-1-carboximidamide:3-(2,3-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1H-indole-1-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-3-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-indole-1-carboximidamide;3-(2,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1H-indole-1-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-3-(3-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-indole-1-carboximidamide;5-chloro-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1H-indole-1-carboximidamide;2-(2,4-dimethylfuran-3-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1H-indole-1-carboximidamide;2-(3,5-dimethylisothiazol-4-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1H-indole-1-carboximidamide;2-(3,5-dimethylisothiazol-4-yl)-3-(4-hydroxyphenyl)-1H-indole-1-carboxamide:5-chloro-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1H-indole-1-carboximidamide:2-(2,4-dimethylfuran-3-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1H-indole-1-carboximidamide;2-(3,5-dimethylisothiazol-4-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-1H-indole-1-carboximidamide:or2-(3,5-dimethylisothiazol-4-yl)-3-(4-hydroxyphenyl)-1H-indole-1-carboxamide

The ERβ ligand may be2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-2,4,5,6-tetrahydrocyclopenta[c]pyrrole-1-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-3-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-5,6-dihydrocyclopenta[b]pyrrole-1(4H)-carboximidamide;3-(2,6-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-5,6-dihydrocyclopenta[b]pyrrole-1(4H)-carboximidamide;3-(2,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-5,6-dihydrocyclopenta[b]pyrrole-1(4H)-carboximidamide; 3-(2-chloro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-5,6-dihydrocyclopenta[b]pyrrole-1(4H)-carboximidamide;3-(2-chloro-6-fluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-5,6-dihydrocyclopenta[b]pyrrole-1(4H)-carboximidamide;3-(2,3-difluoro-4-hydroxyphenyl-2-(3,5-dimethylisoxazol-4-yl)-N-hydroxy-5,6-dihydrocyclopenta[b]pyrrole-1(4H)-carboximidamide;3-(3,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-5,6-dihydrocyclopenta[b]pyrrole-1(4H)-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-3-(4-hydroxyphenyl)-4,5,6,7-tetrahydro-1H-indole-1-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-3-(2-fluoro-4-hydroxyphenyl)-N-hydroxy-4,5,6,7-tetrahydro-1H-indole-1-carboximidamide;3-(2,6-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-4,5,6,7-tetrahydro-1H-indole-1-carboximidamide;3-(2,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-4,5,6,7-tetrahydro-1H-indole-1-carboximidamide:3-(2-chloro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-4,5,6,7-tetrahydro-1H-indole-1-carboximidamide;3-(2,3-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-4,5,6,7-tetrahydro-1H-indole-1-carboximidamide;or3-(3,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-4,5,6,7-tetrahydro-1H-indole-1-carboximidamide.

The ERβ ligand may beN′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-N′,4′-dihydroxy-5-methyl-[1,1′-biphenyl]-3-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide:2-(3,5-dimethylisoxazol-4-yl)-4′-hydroxy-5-propyl-[1,1′-biphenyl]-3-carboxamide:2-(3,5-dimethylisoxazol-4-yl)-4′-hydroxy-5-propyl-[1,1′-biphenyl]-3-carbonitrile;N′4′-dihydroxy-5-methyl-2-(3-methylthiophen-2-yl)-[,1′-biphenyl]-3-carboximidamide;3′,5′-difluoro-N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide:2-bromo-3′,5′-difluoro-N′,4′-dihydroxy-5-propyl-[,1′-biphenyl]-3-carboximidamide:2-(3,5-dimethylisoxazol-4-yl)-3′,5′-difluoro-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;5-bromo-N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-[1,1′-biphenyl]-3-carboximidamide;5-bromo-N′,4′-dihydroxy-2-iodo-[1,1′-biphenyl]-3-carboximidamide;5′-bromo-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″,5′-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:5″-fluoro-4-hydroxy-2″,5′-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboxamide:5-chloro-2-(3,5-dimethylisoxazol-4-yl-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;5-chloro-2-(3,5-dimethylisoxazol-4-yl)-4′-hydroxy-[1,1′-biphenyl]-3-carboxamide;2-(3,5-dimethylisoxazol-4-yl)-3′-fluoro-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;2-(2,4-dimethylfuran-3-yl)-4′-hydroxy-5-propyl-[,1′-biphenyl]-3-carbonitrile:2-(2,4-dimethylfuran-3-yl)-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;2-(2,4-dimethylfuran-3-yl)-4′-hydroxy-5-propyl-[1,1′-biphenyl]-3-carboxamide;N′,4′-dihydroxy-2-iodo-5-propyl-[1,1′-biphenyl]-3-carboximidamide:N′,4′-dihydroxy-2-(4-methylthiophen-3-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide:N′4-dihydroxy-5′-propyl-2″-(trifluoromethoxy)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(2,4-dimethylthiophen-3-yl)-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;N′,4-dihydroxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-((E)-2-cyclopropylvinyl)-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-2-(3-methylbut-2-en-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;N′,4-dihydroxy-3″-methyl-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″-methyl-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″-ethyl-N′,4-dihydroxy-5′-propyl-[1,1,′:2′,1″-terphenyl]-3′-carboximidamide;N′4′-dihydroxy-5-propyl-2-(thiophen-2-yl)-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-5-propyl-2-(quinolin-5-yl)-[1,1′-biphenyl]-3-carboximidamide;3″-chloro-N′,4-dihydroxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4′-dihydroxy-5-propyl-2-(pyridin-3-yl)-[1,1′-biphenyl]-3-carboximidamide;2-(benzofuran-5-yl)-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;4″-chloro-N′,4-dihydroxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4′-dihydroxy-5-propyl-2-(pyridin-4-yl)-[,1′-biphenyl]-3-carboximidamide;N′4′-dihydroxy-2-(1-phenylvinyl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;2-(5-chlorothiophen-2-yl)-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″-methoxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4′-dihydroxy-2-(isoquinolin-6-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;2-(benzofuran-3-yl)-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4′-dihydroxy-2-(4-methylthiophen-3-yl)-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide:2-(2,4-dimethylthiophen-3-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-2-iodo-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2″,5″-difluoro-N′,4-dihydroxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(1,3-dimethyl-1H-pyrol-2-yl)-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;3″,5″-difluoro-N′,4-dihydroxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(2,4-dimethylfuran-3-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;3′-chloro-5′-fluoro-N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;N′-hydroxy-3-(1H-indazol-5-yl)-2-(3-methylthiophen-2-yl)-5-propylbenzimidamide;3′-fluoro-N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;3′-chloro-N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;3′,5′-dichloro-N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-3′-methyl-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;2′-fluoro-N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide:2′,3′-difluoro-N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;2′,5′-difluoro-N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-2-(2-methylallyl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide:2-allyl-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-5-propyl-2-vinyl-[1,1′-biphenyl]-3-carboximidamide;5-bromo-N′,4′-dihydroxy-2-(1-methyl-1H-imidazol-5-yl)-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-5-propyl-2-(pyridin-2-yl)-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-2-(2-methoxythiazol-4-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide:N′,4′-dihydroxy-5-propyl-2-(thiazol-5-yl)-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-5-propyl-2-(thiazol-2-yl)-[1,1′-biphenyl]-3-carboximidamide;5′-ethyl-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-5′-isobutyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-5′-((E)-prop-1-en-1-yl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-allyl-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-butyl-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(2,5-dimethyl-1H-pyrrol-1-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;4-hydroxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carbaldehyde oxime5′-propyl-3′-(1H-pyrazol-4-yl)-[1,1′:2′,1″-terphenyl]-4-ol;5-chloro-2-(3,5-dimethylisoxazol-4-yl)-N′,4′-dihydroxy-3′-nitro-[1,1′-biphenyl]-3-carboximidamide;5′-chloro-5″-fluoro-N′,4-dihydroxy-2″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-5″-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5-chloro-N′,4′-dihydroxy-2-(4-methylthiophen-3-yl)-[1,1′-biphenyl]-3-carboximidamide;5′-chloro-N′,4-dihydroxy-2″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(2,4-dimethylthiophen-3-yl)-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;5-chloro-2-(2,4-dimethylthiophen-3-yl-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;2″-chloro-5″-fluoro-N′,4-dihydroxy-5′-propyl-[,1′:2′,1″-terphenyl]-3′-carboximidamide;6′-chloro-N′,4-dihydroxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-5′,6′-dipropyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:N′,4′-dihydroxy-2-(3-methylthiophen-2-yl)-[1,1′-biphenyl]-3-carboximidamide:5′-bromo-6′-chloro-N′,4-dihydroxy-[,1′:2′,1″-terphenyl]-3′-carboximidamide;6′-chloro-N′,4-dihydroxy-5′-phenyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;6′-chloro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-6′-methyl-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:5″,6′-difluoro-N′,4-dihydroxy-2″-methyl-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″,6′-difluoro-4-hydroxy-2″-methyl-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboxamide;4-hydroxy-6′-methyl-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carbaldehydeoxime5″,6′-difluoro-N′,4-dihydroxy-2″-methoxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″,6′-difluoro-4-hydroxy-2″-methoxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboxamide:6′-fluoro-4-hydroxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboxamide;6′-fluoro-N′,4-dihydroxy-2″-methyl-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;6-fluoro-N′,4′-dihydroxy-2-(4-methylthiophen-3-yl)-5-propyl-[1,1′-biphenyl]-3-carboximidamide;6′-fluoro-4-hydroxy-2″-methyl-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboxamide;6-fluoro-4′-hydroxy-2-(4-methylthiophen-3-yl)-5-propyl-[1,1′-biphenyl]-3-carboxamide;2-(2,4-dimethylthiophen-3-yl)-6-fluoro-N′,4′-dihydroxy-5-propyl-[1,1′-biphenyl]-3-carboximidamide;5′-chloro-5″-fluoro-4-hydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboxamide;6′-fluoro-N′,4-dihydroxy-5′-propyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-5′-(trifluoromethyl)-2″-vinyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″-ethyl-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-fluoro-N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-fluoro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′,6′-dichloro-5″-fluoro-N′4-dihydroxy-2″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5″-difluoro-N′4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-N′,4-dihydroxy-2″-methoxy-5′-(trifluormethyl)-[1,1′:2′,1″-terphenyl]-3-carboximidamide;2″-ethynyl-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-chloro-5″-fluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″,5′-bis(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3′,5″-difluoro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-methyl-3′-(1H-pyrazol-4-yl)-[1,1′:2′,1″-terphenyl]-4-ol;3″,5″-difluoro-5′-propyl-3′-(1H-pyrazol-4-yl)-[1,1′:2′,1″-terphenyl]-4-ol;N′,4-dihydroxy-5′-phenyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-benzyl-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-5′-phenethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2,5″-difluoro-N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2,5″-difluoro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-fluoro-N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-fluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-2-fluoro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2,3″-difluoro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2,3″,5″-trifluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-fluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-2″-vinyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-2-fluoro-4-hydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;2-fluoro-4-hydroxy-5′-(trifluoromethyl)-2″-vinyl-[1,1′:2′,1″-terphenyl]-3′-carboxamide;3″-chloro-2,5″-difluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-chloro-2,5″-difluoro-4-hydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;2″-ethynyl-2-fluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-3″,5″-difluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-3″-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′,5″-dichloro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-chloro-2-fluoro-N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-chloro-2-fluoro-4-hydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;N′,4-dihydroxy-5′-methyl-2″-vinyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:3″,5″-difluoro-N′,4-dihydroxy-2″-methoxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-fluoro-N′,4-dihydroxy-2″-methoxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″-methoxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-chloro-5″-fluoro-N′,4-dihydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-N′,4-dihydroxy-2″,5′-dimethyl-[1,1′:2′,]″-terphenyl)-3′-carboximidamide;5″-chloro-N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-chloro-N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-N′,4-dihydroxy-2″-methoxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″-chloro-5″-fluoro-N′,4-dihydroxy-5′-methyl-[1,1′:2′1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-3″,5′-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-chloro-N′,4-dihydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-3″-methoxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4″-fluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4″-chloro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4″-fluoro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-2″-isopropyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-4′-hydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboxamide;3″-chloro-4-hydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;N′,4-dihydroxy-2″,5″-dimethyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-2″-methoxy-5″-methyl-5′-(trifluoromethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:2″-chlor-N′,4-dihydroxy-5″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-fluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-5′-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5′-difluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5′,5″-trifluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-chloro-5′,5″-difluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-fluoro-N′,4-dihydroxy-2″-vinyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-N′,4-dihydroxy-2″,5″-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:N′,4-dihydroxy-2″,5″-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″-chloro-4-hydroxy-5″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;3″-chloro-N′,4-dihydroxy-5″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″,5″-dichloro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-N′,4-dihydroxy-2″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5″-difluoro-N′,4-dihydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5″-difluoro-4-hydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboxamide;3″-chloro-4-hydroxy-5″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;2″,5″-dichloro-4-hydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;3″,5″-dichloro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5″-dichloro-N′,4-dihydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-2′-fluoro-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-2′-fluoro-4′-hydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboxamide;3″,5″-difluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5″-difluoro-4-hydroxy-[1,1′:2′,1″-terphenyl]-3′-carboxamide:3″-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″-fluoro-4-bydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboxamide:2″-ethyl-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4″-chloro-2″-fluoro-N′,4-dihydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4″-chloro-3″-fluoro-N′,4-dihydroxy-5′-methyl-[,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4″-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-4″-fluoro-N′:4-dihydroxy-2″-methoxy-[1,l″-terphenyl]-3′-carboximidamide;3″-chloro-N′,4-dihydroxy-5′,5″-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:2″,5″-dichloro-N′,4-dihydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5″-difluoro-4-hydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;N′,4-dihydroxy-4″,5′-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-2″,4″,5′-trimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4″-fluoro-N′,4-dihydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″,4″-difluoro-N′4-dihydroxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-5′-fluoro-N′,4-dihydroxy-2″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′,5″-difluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″-chloro-5′-fluoro-N′,4-dihydroxy-5″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″,5″-dichloro-5′-fluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:5′-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4″,5′-difluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-fluoro-N′,4-dihydroxy-2″,5″-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-fluoro-N′4-dihydroxy-2″-methoxy-5″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-2″-methoxy-5″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-2″-methoxy-5′,5″-dimethyl-[,1′:2′,1″-terphenyl]-3′-carboximidamide;4″-fluoro-N′,4-dihydroxy-2″-methoxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:2″-chloro-N′,4-dihydroxy-5′,5″-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4-dihydroxy-5′-methyl-2″-(trifluoromethoxy)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′4-dihydroxy-2″-methoxy-5′-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5″-difluoro-N′,4-dihydroxy-5′-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-fluoro-N′,4-dihydroxy-2″,5′-dimethoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(5-fluoro-2-methoxypyridin-3-yl)-N′,4′-dihydroxy-5-methyl-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-5-methyl-2-(2-methylpyridin-3-yl)-[1,1′-biphenyl]-3-carboximidamide;N′,4′-dihydroxy-2-(2-methoxypyridin-3-yl)-5-methyl-[1,1′-biphenyl]-3-carboximidamide;2-(3,5-dimethylisothiazol-4-yl)-N′,4′-dihydroxy-5-methyl-[1,1′-biphenyl]-3-carboximidamide;N′,4-dihydroxy-5′-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(3,5-dimethylisothiazol-4-yl)-4′-hydroxy-5-methyl-[1,1′-biphenyl]-3-carboxamide;5-chloro-2-(5-fluoro-2-methoxypyridin-3-yl)-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;2-(5-fluoro-2-methoxypyridin-3-yl)-4′-hydroxy-5-methyl-[1,1′-biphenyl]-3-carboxamide;2-(3,5-dimethylisothiazol-4-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;5-chloro-2-(3,5-dimethylisothiazol-4-yl)-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;4″-fluoro-N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″,4″-difluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4′-dihydroxy-2-(4-methylpyridin-3-yl)-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2-(2,5-dimethylpyridin-3-yl)-N′,4′-dihydroxy-5-(trifluormethyl)-[1,1′-biphenyl]-3-carboximidamide;2″-chloro-4″-fluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4″,5″-difluoro-N′,4-dihydroxy-2″-methyl-5′-(trifluoromethyl)-)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″,5″-difluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;N′,4′-dihydroxy-2-(pyridin-3-yl)-5-(trifluormethyl)-[1,1′-biphenyl]-3-carboximidamide;2-(2,3-dihydrobenzofuran-7-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide:2-(benzofuran-7-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;N′4′-dihydroxy-2-(1-methyl-1H-indol-7-yl)-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2″-chloro-5″-fluoro-N′,4-dihydroxy-5′-(trifluromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-2″-fluoro-N′,4-dihydroxy-5′-(trifluromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(4-fluorobenzofuran-7-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;3″,4″-difluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-2″,4″-difluoro-N′,4-dihydroxy-[1,1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-2″,5″-difluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(benzo[d][1,3]dioxol-4-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;3″,4″,5″-trifluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-3″,4″-difluoro-N′,4-dihydroxy-[1,1′:2′1″-terphenyl]-3′-carboximidamide;5′-chloro-3″,4″,5″-trifluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5-chloro-2)N′,4′-dihydroxy-2-(4-methylpyridin-3-yl)-[1,1′-biphenyl]-3-carboximidamide;5′,5″-dichloro-2″-fluoro-N′,4-dihydroxy-[1,1′:2′1″-terphenyl]-3′-carboximidamide; 2″,5dichloro-5″-fluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-2″-fluoro-N′,4-dihydroxy-5″-methyl-[,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-2″-fluoro-N′,4-dihydroxy-4″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(5-fluoro-2-methoxypyridin-3-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;5′-chloro-N′,4-dihydroxy-2″,4″-dimethoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-4-hydroxy-2″,4″-dimethoxy-[1,1′:2′,1″-terphenyl]-3′-carbonitrile;5-chloro-2-(5-chloro-2-methoxypyridin-3-yl)-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;5-chloro-2-(5-chloro-2-methoxypyridin-3-yl)-4′-hydroxy-[1,1′-biphenyl]-3-carboxamide;5-chloro-2-(2,5-dimethylpyridin-3-yl)-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;5-chloro-2-(2,5-dimethylpyridin-3-yl)-4′-hydroxy-[1,1′-biphenyl]-3-carboxamide:2-(benzo[d][1,3]dioxol-4-yl)-5-chloro-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;5-chloro-N′,4′-dihydroxy-2-(naphthalen-2-yl)-[1,1′-biphenyl]-3-carboximidamide;5-chloro-N′,4′-dihydroxy-2-(isoquinolin-6-yl)-[1,1′-biphenyl]-3-carboximidamide;5-chloro-N′,4′-dihydroxy-2-(quinolin-6-yl)-[1,1′-biphenyl]-3-carboximidamide:5-chloro-N′,4′-dihydroxy-2-(1-methyl-H-benzo[d]imidazol-5-yl)-[1,1′-biphenyl]-3-carboximidamide;5-chloro-4′-hydroxy-2-(1-methyl-1H-benzo[d]imidazol-5-yl)-[1,1′-biphenyl]-3-carboxamide;2-(5-chloro-2-methoxypyridin-3-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2″-fluoro-N′,4-dihydroxy-5″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5-chloro-2-(6-chloro-2-methoxypyridin-3-yl)-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;5-chloro-N′,4′-dihydroxy-2-(2-methoxy-5-methylpyridin-3-yl)-[1,1′-biphenyl]-3-carboximidamide;5-chloro-2-(cyclopent-1-en-1-yl)-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;5-chloro-2-(cyclopent-1-en-1-yl)-4′-hydroxy-[1,1′-biphenyl]-3-carboxamide;2-(cyclopent-1-en-1-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2-(cyclopent-1-en-1-yl)-4′-hydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboxamide;5′-bromo-5″-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-5″-fluoro-N′,4-dihydroxy-2″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-3″-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-N′,4-dihydroxy-2″,5″-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-N′,4-dihydroxy-5″-methoxy-2″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-4″-fluoro-N′,4-dihydroxy-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-fluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-2″-vinyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2,5″-difluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-2″-vinyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-N′,4-dihydroxy-5′-(trifluoromethyl)-2″-vinyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2″-ethynyl-5″-fluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4-hydroxy-2″-methoxy-5″-methyl-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;5′-bromo-5″-chloro-N′,4-dihydroxy-2″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-3″,5″-difluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-3″-chloro-5″-fluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:5′-bromo-2″-chloro-5″-fluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:5′-bromo-4″-chloro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-2″,5″-dichloro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-bromo-2″-chloro-N′,4-dihydroxy-5″-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5-bromo-2-(3,5-dimethylisoxazol-4-yl)-N′,4′-dihydroxy-[1,1′-biphenyl]-3-carboximidamide;3-chloro-5-fluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3-chloro-5-fluoro-4-hydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;3,5′-dichloro-3″,5,5″-trifluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3′-chloro-2-(3,5-dimethylisoxazol-4-yl)-5′-fluoro-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;3-chloro-3″,5,5″-trifluoro-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:3-chloro-5,5″-difluoro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3-chloro-5-fluoro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethyl)-[1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-3″,5″-difluoro-N′-hydroxy-4-methoxy-3-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-3″,5″-difluoro-N′,4-dihydroxy-3-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5-chloro-2-(3,5-dimethylisoxazol-4-yl)-N′,4′-dihydroxy-3′-methyl-[1,1′-biphenyl]-3-carboximidamide;5′-chloro-N′,4-dihydroxy-2″,3-dimethyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-5″-fluoro-N′,4-dihydroxy-2″-methoxy-3-methyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5′-chloro-5″-fluoro-4-hydroxy-2″-methoxy-3-methyl-[1,1′:2′1″-terphenyl]-3′-carboxamide;5′-chloro-3″,5″-difluoro-N′,4-dihydroxy-3-isopropyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2′-(3,5-dimethylisoxazol-4-yl)-3′-(1H-1,2,3-triazol-4-yl)-5′-(trifluoromethyl)-[1,1′-biphenyl]-4-ol;N′,4-dihydroxy-5′-(trifluoromethoxy)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5″-difluoro-N′,4-dihydroxy-5′-(trifluoromethoxy)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-N′,4-dihydroxy-2″-methoxy-5′-(trifluoromethoxy)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:4-hydroxy-5′-(trifluoromethoxy)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;3″,5″-difluoro-4-hydroxy-5′-(trifluoromethoxy)-[1,1′:2′,1″-terphenyl]-3′-carboxamide;N′,4-dihydroxy-5′-isopropyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;3″,5″-difluoro-N′,4-dihydroxy-5′-isopropyl-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;5″-chloro-N′,4-dihydroxy-5′-isopropyl-2″-methoxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;4-amino-5′-isopropyl-[1,1′:2′,1″-terphenyl]-3′-carboxamide;3″,5″-difluoro-4-hydroxy-5′-isopropyl-[1,1′:2′,1″-terphenyl]-3′-carboxamide;2-(3-cyanofuran-2-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2-(4-cyano-1-methyl-1H-pyrazol-5-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2″-cyano-N′,4-dihydroxy-5′-(trifluoromethyl)-[1,1′:2′,1″-terphenyl]-3′-carboximidamide;2-(3-cyano-1-methyl-1H-pyrrol-2-yl)-N′,4′-dihydroxy-5-(trifluoromethyl)-[1,1′-biphenyl]-3-carboximidamide;2′-(3,5-dimethylisoxazol-4-yl)-3′-(hydroxymethyl)-5′-(trifluoromethyl)-[1,1′-biphenyl]-4-ol;5′-cyano-3″,5″-difluoro-N′,4-dihydroxy-[1,1′:2′,1″-terphenyl]-3′-carboximidamide:N′,4′-dihydroxy-2-(pyrrolidin-1-yl)-5-(trifluoromethyl)[1,1′-biphenyl]-3-carboximidamide;4-chloro-3′,5′-difluoro-N′-hydroxy-6-(1H-indazol-5-yl)-[1,1′-biphenyl]-2-carboximidamide;6-(1H-benzo[d]imidazol-5-yl)-4-chloro-N′-hydroxy-[1,1′-biphenyl]-2-carboximidamide;4-chloro-3′,5′-difluoro-N′-hydroxy-6-(1H-indol-5-yl)-[1,1′-biphenyl]-2-carboximidamide;4-chloro-3′,5′-difluoro-6-(1H-indol-5-yl)-[,1′-biphenyl]-2-carboxamide;4-chloro-3′,5′-difluoro-6-(1H-indazol-5-yl)-[1,1′-biphenyl]-2-carboxamide;6-(1H-benzo[d]imidazol-5-yl)-4-chloro-3′,5′-difluoro-[1,1′-biphenyl]-2-carboxamide;3′,5-difluoro-N′-hydroxy-6-(1H-indol-5-yl)-4-methyl-[1,1′-biphenyl]-2-carboximidamide;3′,5′-difluoro-N′-hydroxy-6-(1H-indol-6-yl)-4-methyl-[1,1′-biphenyl]-2-carboximidamide;3′,5′-difluoro-N′-hydroxy-6-(1H-indazol-5-yl)-4-methyl-[1,1′-biphenyl]-2-carboximidamide;3′,5′-difluoro-N′-hydroxy-6-(1H-indazol-5-yl)-4-propyl-[1,1′-biphenyl]-2-carboximidamide;3′,5′-difluoro-N′-hydroxy-6-(1H-indol-5-yl)-4-propyl-[1,1′-biphenyl]-2-carboximidamide;3′5′-difluoro-N′-hydroxy-6-(1H-indazol-5-yl)-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;3′,5′-difluoro-N′-hydroxy-6-(1H-indol-6-yl)-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide:3′,5′-difluoro-N′-hydroxy-6-(1H-indazol-6-yl)-4-(trifluoromethyl)-[1,16′-biphenyl]-2-carboximidamide;3′,5′-difluoro-N′-hydroxy-6-(1H-indol-5-yl)-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;3′,5′-difluoro-6-(1H-indol-5-yl)-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboxamide;3′,5′-difluoro-N′-hydroxy-6-(1H-indazol-6-yl)-4-propyl-[1,1′-biphenyl]-2-carboximidamide;N′-hydroxy-6-(1H-indazol-5-yl)-2′-methoxy-5′-methyl-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;5′-chloro-N′-hydroxy-6-(1H-indol-5-yl)-2′-methoxy-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide:5′-chloro-N′-hydroxy-6-(1H-indazol-5-yl)-2′-methoxy-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;2′,5′-difluoro-N′-hydroxy-6-(1H-indazol-5-yl)-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;3′,5′-difluoro-6-(6-fluoro-1H-indol-5-yl)-N′-hydroxy-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;N′-hydroxy-3-(1H-indazol-5-yl)-2-(naphthalen-1-yl)-5-(trifluoromethyl)benzimidamide;2-(benzo[d][1,3]dioxol-4-yl)-N′-hydroxy-3-(IH-indazol-5-yl)-5-(trifluoromethyl)benzimidamide;4′-fluoro-N′-hydroxy-6-(1H-indazol-5-yl)-2′-methoxy-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;N′-hydroxy-6-(IH-indazol-5-yl)-3′-methyl-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;3′,4′,5′-trifluoro-N′-hydroxy-6-(1H-indazol-5-yl)-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;3′,4′-difluoro-N′-hydroxy-6-(1H-indazol-5-yl)-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide;N′-hydroxy-3-(1H-indazol-5-yl)-2-(2-methoxypyridin-3-yl)-5-(trifluoromethyl)benzimidamide;2-(cyclopent-1-en-1-yl)-N′-hydroxy-3-(1H-indazol-5-yl)-5-(trifluoromethyl)benzimidamide;2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-3-(IH-indazol-5-yl)-5-(trifluoromethyl)benzimidamide; or2′,4′-difluoro-N′-hydroxy-6-(1H-indazol-5-yl)-4-(trifluoromethyl)-[1,1′-biphenyl]-2-carboximidamide.

The ERβ ligand may be2-Bromo-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-hydroxy-phenyl)-2-thiophen-3-yl-1H-indole-3-carbonitrile;2-(3-Cyano-furan-2-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-pyrrol-1-yl-1H-indole-3-carbonitrile;2-Dimethylamino-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-isopropyl-1H-indole-3-carbonitrile;2-Acetyl-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-(3,5-Dimethyl-isoxazol-4-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carboxylicacid; I-[l-(4-Hydroxy-phenyl)-2-phenyl-1H-indol-3-yl]-ethanone;1-(4-Hydroxy-phenyl)-2-phenyl-1H-indole-3-carboxylic acid, amide:(Z)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;(2-(3,5-Dimethyl-isoxazol-4-yl)-1-(4-hydroxy-phenyl)-1H-indol-3-yl-carbamicacid tert-butyl ester;4-[3-Amino-2-(3,5-dimethyl-isoxazol-4-yl)-indol-1-yl]-phenol;(Z)-2-(3,5-dimethylisoxazol-4-yl)-7-fluoro-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;(Z)-2-(5-chlorothiophen-2-yl)-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-(3,5-dimethyl-isoxazol-4-yl)-1H-indole-3-carbonitrile;2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indole-3-carbohydrazonamide;4-(2-(3,5-dimethylisoxazol-4-yl)-3-(1,2,4-oxadiazol-3-yl)-1H-indol-1-yl)phenol;2-(3,5-Dimethyl-isoxazol-4-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carboxylicacid, methyl ester;2-(3,5-Dimethyl-isoxazol-4-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carboxylicacid, hydroxyamide;4-[2-(3,5-Dimethyl-isoxazol-4-yl)-3-methanesulfonyl-indol-1-yl]-phenol;1-[2-(3,5-Dimethyl-isoxazol-4-yl)-1-(4-hydroxy-phenyl)-1H-indol-3-yl]-2,2,2-trifluoro-ethanone;4-(3-bromo-2-(3,5-dimethylisoxazol-4-yl)-1H-indol-1-yl)phenol;2-Bromo-5-fluoro-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;(Z)-2-(4-fluorophenoxy)-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indol-3-carboximidamide:4-(2-(3,5-dimethylisoxazol-4-yl)-3-nitro-1H-indol-1-yl)phenol4-(3-(dihydroxyamino)-2-(3,5-dimethylisoxazol-4-yl)-1H-indol-1-yl)phenol;N-(2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indol-3-yl)acetamideN-(2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indol-3-yl)methanesulfonamide;1-(2,3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indol-3-yl)urea;4-(2-(3,5-dimethylisoxazol-4-yl)-3-thiocyanato-1H-indol-1-yl)phenol;(E)-2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indol-3-ylN′-hydroxycarbamimidothioate; 4-(3-benzyl-2-phenyl-1H-indol-1-yl)phenol;2-(2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indol-3-yl)-2-oxoacetamide;(Z)-2-(2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indol-3-yl)-2-(hydroxyimino)acetamide;2-(2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indol-3-yl)-2-hydroxyacetamide;2-(2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indol-3-yl)acetamide;2-((Z)-But-1-enyl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(2-methyl-prop-1-enyl)-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-(2-methyl-allyl)-1H-indole-3-1acarbonitrile;(Z)-2-(5-ethyl-3-methylisoxazol-4-yl)-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;4-(2-(3,5-dimethylisoxazol-4-yl)-3-phenyl-1H-indol-1-yl)phenol;4-(3-chloro-2-(3,5-dimethylisoxazol-4-yl)-1H-indol-1-yl)phenol;2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indole-3-sulfonamide;2-(3,5-dimethylisoxazol-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-1H-indole-3-carboximidamide;1-(4-Hydroxy-phenyl)-2-phenyl-1H-indole-3-carbonitrile;1-(4-hydroxy-phenyl)-2-methyl-1H-indole-3-carbonitrile;2-(3-Cyano-thiophen-2-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-((E)-propenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-thiophen-2-yl-1H-indole-3-carbonitrile;2-(3,5-Dimethyl-isoxazol-4-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-pyridin-4-yl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(1-methyl-1H-pyrrol-2-yl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(3-methyl-thiophen-2-yl)-1H-indole-3-carbonitrile;I-(4-Hydroxy-phenyl)-2-isopropylamino-1H-indole-3-carbonitrile;2-Ethylamino-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-Butylamino-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-piperidin-1-yl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-pyrrolidin-1-yl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-morpholin-4-yl-1H-indole-3-carbonitrile;2-Diethylamino-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-Ethynyl-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-vinyl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-H-indol-2,3-dicarbonitrile:1-(4-Hydroxy-phenyl)-2-prop-1-ynyl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-pyridin-2-yl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(2-methyl-allyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-((Z)-propenyl)-1H-indole-3-carbonitrile;2-(Butyl-methyl-amino)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-((Z)-1-methyl-propenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-imidazol-1-yl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-[1,2,4]triazol-1-yl-1H-indole-3-carbonitrile;2-(3,5-Dimethyl-pyrazol-1-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-pyrazol-1-yl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(5-methyl-imidazol-1-yl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(5-methyl-pyrazol-1-yl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(3-methyl-pyrazol-1-yl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-thiazol-2-yl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(2-methoxy-thiazol-4-yl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-thiazol-4-yl-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(3-methyl-but-2-enyl)-1H-indole-3-carbonitrile;2-((E)-But-1-enyl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(5-methyl-thiophen-2-yl)-1H-indole-3-carbonitrile;2-(5-Acetyl-thiophen-2-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(I-methyl-1H-pyrazol-4-yl)-1H-indole-3-carbonitrile;2-(5-Chloro-thiophen-2-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;I-(4-Hydroxy-phenyl)-2-(4-methyl-thiophen-3-yl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(4-methyl-thiophen-2-yl)-1H-indole-3-carbonitrile;2-(4-Cyano-thiophen-3-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydoxy-phenyl)-2-(2-methyl-2H-pyrazol-3-yl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(1,3,5-trimethyl-1H-pyrazol-4-yl)-1H-indole-3-carbonitrile;2-(2-Acetyl-pyrrol-1-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-(2-Ethyl-pyrrol-1-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-21,carbonitrile;2-(2-Cyano-pyrrol-1-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(4-Hydroxy-phenyl)-2-(2-methyl-pyrrol-2-yl)-1H-indole-3-carbonitrile;1-(3-Chloro-5-fluoro-4-hydroxy-phenyl)-2-phenyl-1H-indole-3-carbonitrile;1-(3-Chloro-5-fluoro-4-hydroxy-phenyl)-2-(3-cyano-thiophen-2-yl)-1H-indole-3-carbonitrile;1-(3-Chloro-5-fluoro-4-hydroxy-phenyl)-2-(3-cyano-furan-2-yl)-1H-indole-3-carbonitrile;2-Bromo-1-(3-chloro-5-fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-Bromo-1-(2-fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(2-Fluoro-4-hydroxy-phenyl)-2-phenyl-1H-indole-3-carbonitrile;2-Bromo-1-(3-fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-Bromo-1-(2,3-difluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-Bromo-1-(2,5-difluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-Bromo-1-(3-chloro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-Bromo-1-(3,5-difluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-phenyl-1H-indole-3-carbonitrile;1-(3,5-Difluoro-4-hydroxy-phenyl)-2-phenyl-1H-indole-3-carbonitrile;1-(3-Chloro-4-hydroxy-plenyl)-2-phenyl-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-phenyl-1H-indole-3-carbonitrile;1-(2,5-Difluoro-4-hydroxy-phenyl)-2-phenyl-1H-indole-3-carbonitrile;1-(3,5-Difluoro-4-hydroxy-phenyl)-2-thiophen-3-yl-1H-indole-3-carbonitrile;1-(3,5-Difluoro-4-hydroxy-phenyl)-2-thiophen-2-yl-1H-indole-3-carbonitrile;1-(3,5-Difluoro-4-hydroxy-phenyl)-2-(3,5-dimethyl-isoxazol-4-yl)-1H-indole-3-carbonitrile;1-(3,5-Difluoro-4-hydroxy-phenyl)-2-(1-methyl-1H-pyrrol-2-yl)-1H-indole-3-carbonitrile;1-(3,5-Difluoro-4-hydroxy-phenyl)-2-(3-methyl-thiophen-2-yl)-1H-indole-3-carbonitrile;1-(3,5-Difluoro-4-hydroxy-phenyl)-2-(1-methyl-1H-pyrazol-4-yl)-1H-indole-3-carbonitrile;1-(3,5-Difluoro-4-hydroxy-phenyl)-2-pyridin-4-yl-H-indole-3-carbonitrile;1H-(3-Chloro-4-hydroxy-phenyl)-2-thiophen-3-yl-1H-1-indole-3-carbonitrile;1-(3-Chloro-4-hydroxy-phenyl)-2-thiophen-2-yl-1H-indole-3-carbonitrile;1-(3-Chloro-4-hydroxy-phenyl)-2-(3,5-dimethyl-isoxazol-4-yl)-1H-indole-3-carbonitrile;1-(3-Chloro-4-hydroxy-phenyl)-2-(1-methyl-1H-pyrrol-2-yl)-1H-indole-3-carbonitrile;1-(3-Chloro-4-hydroxy-phenyl)-2-(3-methyl-thiophen-2-yl-1H-indole-3-carbonitrile;1-(3-Chloro-4-hydroxy-phenyl)-2-pyridin-4-yl-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-thiophen-3-yl-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-thiophen-2-yl-1H-indole-3-carbonitrile;2-(3,5-Dimethyl-isoxazol-4-yl)-1-(3-Fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-(1-methyl-1H-pyrrol-2-yl)-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-(3-methyl-thiophen-2-yl)-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-(1-methyl-1H-pyrazol-4-yl)-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-pyridin-4-yl-1H-indole-3-carbonitrile;2-Dimethylamino-1-(2-fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-(3,5-Dimethyl-isoxazol-4-yl)-1-(2-fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;i-(3-Fluoro-4-hydroxy-phenyl)-2-((E)-propenyl)-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-((Z)-propenyl)-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-((Z)-propenyl)-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-vinyl-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-thiophen-3-yl-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-thiophen-2-yl-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-(3-methyl-thiophen-2-yl)-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-(1-methyl-1H-pyrrol-2-yl)-1H-indole-3-carbonitrile;2-(2-Acetyl-pyrrol-1-yl)-1-(3-fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-pyrrol-1-yl-1H-indole-3-carbonitrile:1-(2,3-Difluoro-4-hydroxy-phenyl)-2-pyrrol-1-yl-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-prop-1-ynyl-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-(2-methyl-prop-1-enyl)-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-(2-methyl-prop-1-enyl)-1H-indole-3-carbonitrile;2-(2-Acetyl-pyrrol-1-yl)-1-(2,3-difluoro-4-hydroxy-phenyl)-1-H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-pyrazol-1-yl-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-pyrazol-1-yl-H-1-indole-3-carbonitrile;2-(2,5-Dimethyl-pyrrol-1-yl)-1-(3-fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-(2-Ethyl-pyrrol-1-yl)-1-(3-fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-(2-Cyano-pyrrol-1-yl)-1-(3-fluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(3-Fluoro-4-hydroxy-phenyl)-2-(2-methyl-pyrrol-1-yl)-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-(2-ethyl-pyrrol-1-yl)-1H-indole-3-carbonitrile;2-(2-Cyano-pyrrol-1-yl)-1-(2,3-difluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-(2-methyl-pyrrol-1-yl)-1H-indole-3-carbonitrile;1-(2-Fluoro-4-hydroxy-phenyl)-2-pyrrol-1-yl-1H-indole-3-carbonitrile:1-(2,3-difluoro-4-hydroxyphenyl)-2-(3-methylbut-2-enyl)-1H-indole-3-carbonitrile;[1-(4-Hydroxy-phenyl)-2-phenyl-1H-indol-3-yl]-acetonitrile;[1-(4-Hydroxy-phenyl)-2-phenyl-1H-indol-3-yl]-acetic acid;2-[1-(4-Hydroxy-phenyl)-2-phenyl-1H-indol-3-yl]-acetamide:4-(3-Isopropenyl-2-phenyl-indol-1-yl)-phenol;4-[3-(2-Methyl-2H-pyrazol-3-yl)-2-phenyl-indo-1-yl]-phenol;4-(2-Phenyl-3-thiazol-4-yl-indol-1-yl)-phenol;4-(2-Phenyl-3-prop-1-ynyl-indol-1-yl)-phenol;1-(4-Hydroxy-phenyl)-2-((E)-propenyl)-1H-indole-3-carboxylic acid,amide:1-(4-Hydroxy-phenyl)-2-(2-methyl-prop-1-enyl)-1H-indole-3-carboxylicacid, amide;1-(4-Hydroxy-phenyl)-2-((Z)-1-methyl-propenyl)-1H-indole-3-carboxylicacid,amide; 4-(2-Phenyl-3-pyrazol-1-yl-indol-1-yl)-phenol;4-(3-Imidazol-1-yl-2-phenyl-indol-1-yl)-phenol;4-[3-(5-Methyl-pyrazol-1-yl)-2-phenyl-indol-1-yl]-phenol;2-Bromo-1-(4-hydroxy-phenyl)-1H-indole-3-carboxylic acid, amide;1-(4-Hydroxy-phenyl)-2-((Z)-3,3,3-trifluoro-propenyl)-1H-indole-3-carbonitrile;(Z)-2-bromo-N′-hydroxy-1-(4-hydroxyphenyl)-H-indole-3-carboxinidamide;(Z)—N′-hydroxy-1-(4-hydroxyphenyl)-2-(1H-pyrrol-1-yl)-1H-indole-3-carboximidamide;2-(3,5-Dimethyl-isoxazol-4-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carboxylicacid, amide;(Z)—N′-hydroxy-1-(4-hydroxyphenyl)-2-(2-methylprop-1-enyl)-1H-indole-3-carboximidamide;1-(4-Hydroxy-phenyl)-2-phenyl-1H-indole-3-carboxylic acid; hydroxyamide;(Z)—N′-hydroxy-1-(4-hydroxyphenyl)-2-phenyl-1H-indole-3-carboximidamide;1-(4-Hydroxy-phenyl)-2-pyrrol-1-yl-1H-indole-3-carboxylic acid, amide;[1-(4-Hydroxy-phenyl)-2-pyrrol-1-yl-1H-indol-3-yl]-carbamic acidtert-butyl ester;2-(3,5-Dimethyl-isoxazol-4-yl)-1-(4-hydroxy-phenyl)-N-methyl-1H-indole-3-carboxamidine;Methyl2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indole-3-carbimidate;N-((2-(3,5-dimethylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indol-3-yl)(imino)methyl)acetamide;2-(5-ethyl-3-methylisoxazol-4-yl)-1-(4-hydroxyphenyl)-1H-indole-3-carboxamide;(Z)-2-(2-ethyl-1H-pyrrol-1-yl)-N-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;(Z)—N′-hydroxy-1-(4-hydroxyphenyl)-2-(2-methyl-1H-pyrrol-1-yl)-1H-indole-3-carboximidamide;1-(4-hydroxyphenyl)-2-(2-methyl-1H-pyrrol-1-yl)-1H-indole-3-carboxamide;4-(3-chloro-2-(3,5-dimethylisoxazol-4-yl)-1H-indol-1-yl)phenol;(Z)-2-((Z)-but-2-en-2-yl)-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;(Z)—N′-hydroxy-1-(4-hydroxyphenyl)-2-(5-methyl-1H-pyrazol-1-yl)-1H-indole-3-carboximidamide;(Z)—N′-hydroxy-1-(4-hydroxyphenyl)-2-(4-methylthiophen-3-yl)-1H-indole-3-carboximidamide;(Z)-2-(2,5-dimethyl-1H-pyrrol-1-yl)-1H-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;(Z)—N′-hydroxy-1-(4-hydroxyphenyl)-2-phenoxy-1H-indole-3-carboximidamide;1-(4-Hydroxy-phenyl)-2-phenyl-1H-indole-3-carboxylic acid;2-(3,5-)methyl-isoxazol-4-yl)-1-(3-fluoro-4-hydroxy-phenyl)-1H-indole-3-carboxylicacid;2-(3,5-Dimethyl-isoxazol-4-yl)-1-(2-fluoro-4-hydroxy-phenyl)-H-indole-3-carboxylicacid;1-(2,3-Difluoro-4-hydroxy-phenyl)-2-(3,5-dimethyl-isoxazol-4-yl)-1H-indole-3-carboxylicacid; 1-(4-Hydroxy-phenyl)-2-((Z)-propenyl)-1H-indole-3-carboxylic acid;1-(4-Hydroxy-phenyl)-2-((E)-propenyl)-1H-indole-3-carboxylic acid;1-(4-Hydroxy-phenyl)-2-(2-methyl-prop-1-enyl)-1H-indole-3-carboxylicacid; 1-(4-Hydroxy-phenyl)-2-(2-methyl-allyl)-1H-indole-3-carboxylicacid;1-(4-Hydroxy-phenyl)-2-((Z)-1-methyl-propenyl)-1H-indole-3-carboxylicacid;1-(3-Fluoro-4-hydroxy-phenyl)-2-thiophen-3-yl-1H-indole-3-carboxylicacid;1-(3-Fluoro-4-hydroxy-phenyl)-2-thiophen-2-yl-1H-indole-3-carboxylicacid;1-(3-Fluoro-4-hydroxy-phenyl)-2-(1-methyl-1H-pyrrol-2-yl)-1-indole-3-carboxylicacid;1-(3-Fluoro-4-hydroxy-phenyl)-2-(3-methyl-thiophen-2-yl)-1H-indole-3-carboxylicacid; 2-Bromo-1-(4-hydroxy-phenyl)-1H-indole-3-carboxylic acid:1-(4-Hydroxy-phenyl)-2-pyrrol-1-yl-1H-indole-3-carboxylic acid;2,7-Dibromo-1-(2,5-difluoro-4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-Bromo-4-fluoro-1-(4-hydroxy-phenyl)-1-indole-3-carbonitrile;4-Fluoro-1-(4-hydroxy-phenyl)-2-pyrrol-1-yl-1H-indole-3-carbonitrile;4-Fluoro-1-(4-hydroxy-phenyl)-2-phenyl-1H-indole-3-carbonitrile;2-(3,5-Dimethyl-isoxazol-4-yl)-4-fluoro-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;4-Fluoro-1-(4-hydroxy-phenyl)-2-(2-methyl-prop-1-enyl)-1H-indole-3-carbonitrile;5-Fluoro-1-(4-hydroxy-phenyl)-2-phenyl-1H-indole-3-carbonitrile;2-(3,5-Dimethyl-isoxazol-4-yl)-5-fluoro-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;5-Fluoro-1-(4-hydroxy-phenyl)-2-pyrrol-1-yl-1H-indole-3-carbonitrile;5-Fluoro-1-(4-hydroxy-phenyl)-2-(2-methyl-prop-1-enyl)-1H-indole-3-carbonitrile;(Z)-2-(3,5-dimethylisoxazol-4-yl)-5-fluoro-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide:(Z)-2-(3,5-dimethylisoxazol-4-yl)-4-fluoro-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;(Z)-5-fluoro-N′-hydroxy-1-(4-hydroxyphenyl)-2-(2-methylprop-1-enyl)-1H-indole-3-carboximidamide;4-Chloro-2-(3,5-dimethyl-isoxazol-4-yl)-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-(3,5-dimethylisoxazol-4-yl)-4,5-difluoro-1-(4-hydroxyphenyl)-1H-indole-3-carbonitrile;2-(4-cyano-1-methyl-1H-pyrazol-5-yl)-4-fluoro-1-(4-hydroxyphenyl)-1H-indole-3-carbonitrile;2-(3,5-dimethylisoxazol-41-yl)-5-fluoro-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-5-fluoro-1-(4-hydroxyphenyl)-1H-indole-3-carboxamide;2-Bromo-7-fluoro-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-(3,5-Dimethyl-isoxazol-4-yl)-7-fluoro-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile;2-(3,5-Dimethyl-isoxazol-4-yl)-4,7-difluoro-1-(4-hydroxy-phenyl)-1H-indole-3-carbonitrile:(Z)-2-(3,5-dimethylisoxazol-4-yl-4,7-difluoro-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide;1-(2,5-Difluoro-4-hydroxy-phenyl)-2-(3,5-dimethyl-isoxazol-4-yl)-1H-indole-3-carbonitrile;1-(3-bromo-4-hydroxyphenyl)-2-(2-methylprop-1-enyl)-1H-indole-3-carboxamide,(Z)-2-(3,5-dimethylisoxazol-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-indole-3-carboximidamide;(Z)-1-(2,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1H-indole-3-carboximidamide;(Z)-1-(3,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1H-indole-3-carboximidamide;(Z)-2-(3,5-dimethylisoxazol-4-yl)-1-(3-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-indole-3-carboximidamide:(Z)-1-(3-chloro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1H-indole-3-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-1H-indole-3-carboxamide;(Z)-1-(2,3-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-N′-hydroxy-1H-indole-3-carboximidamide;1-(2,3-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-1H-indole-3-carboxamide;1-(2-fluoro-4-hydroxyphenyl)-2-(3-methylthiophen-2-yl)-1H-indole-3-carbonitrile;2-(3,5-dimethyl-1H-pyrazol-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-1H-indole-3-carbonitrile;1-(2-fluoro-4-hydroxyphenyl)-2-(1-methyl-1H-pyrazol-5-yl)-1H-indole-3-carbonitrile;1-(2-fluoro-4-hydroxyphenyl)-2-(1,3,5-trimethyl-1H-pyrazol-4-yl)-1H-indole-3-carbonitrile;1-(2-fluoro-4-hydroxyphenyl)-2-(3-(trifluoromethyl)-1H-pyrazol-4-yl)-1H-indole-3-carbonitrile;(Z)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(3-methylthiophen-2-yl)-1H-indole-3-carboximidamide;(Z)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(1-methyl-1H-pyrazol-5-yl)-1H-indole-3-carboxinidamide;(Z)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(1,3,5-trimethyl-1H-pyrazol-4-yl)-1H-indole-3-carboximidamide;(Z)-4-fluoro-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(I-methyl-H-pyrazol-5-yl)-1H-indole-3-carboximidamide;(Z)-4-fluoro-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(1,3,5-trimethyl-1H-pyrazol-4-yl)-1H-indole-3-carboximidamide;(Z)-2-(3,5-dimethyl-1H-pyrazol-4-yl)-4-fluoro-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-indole-3-carboximidamide;(Z)-4-fluoro-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(3-methylthiophen-2-yl)-1H-indole-3-carboximidamide;(Z)-2-(3,5-dimethyl isoxazol-4-yl)-4-fluoro-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-indole-3-carboximidamide;(Z)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(3-methylthiophen-2-yl)-1H-indole-3-19carboximidamide:(Z)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(1-methyl-1H-pyrazol-5-yl)-1H-indole-3-carboximidamide;(Z)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(1,3,5-trimethyl-1H-pyrazol-4-yl)-1H-indole-3-carboximidamide;(Z)-2-(3,5-dimethyl-1H-pyrazol-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-indole-3-carboximidamide;methyl2-(3,5-dimethylisoxazol-4-yl)-1-(2-fluoro-4-hydroxyphenyl)-1H-indole-3-carbimidate;2-(3,5-dimethylisoxazol-4-yl)-1-(3-fluoro-4-hydroxyphenyl)-1H-indole-3-carboxamide;1-(2,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-1H-indole-3-carboximidamide;2-(3,5-dimethylisoxazol-4-yl)-1-(3-fluoro-4-hydroxyphenyl)-1H-indole-3-carboximidamide;(Z)-1-(3-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(1H-pyrrol-1-yl)-1H-indole-3-carboximidamide;1-(3-fluoro-4-hydroxyphenyl)-2-(1H-pyrrol-1-yl)-H-indole-3-carboxamide;(Z)-1-(2,3-difluoro-4-hydroxyphenyl)-N′-hydroxy-2-(1H-pyrrol-1-yl)-1H-indole-3-carboximidamide;1-(2,3-difluoro-4-hydroxyphenyl)-2-(1H-pyrrol-1-yl)-1H-indol-3-carboxamide;(Z)-2-(2,5-dimethyl-1H-pyrrol-1-yl)-1-(3-fluoro-4-hydroxyphenyl)-N′-hydroxy-1H-indole-3-carboximidamide;(Z)-1-(3-fluoro-4-hydroxyphenyl)-N′-hydroxy-2-(2-methyl-1H-pyrrol-1-yl)-H-indole-3-carboximidamide;1-(3,5-difluoro-4-hydroxyphenyl)-2-(3,5-dimethylisoxazol-4-yl)-1H-indole-3-carboxamide;or(Z)-2-(3,5-dimethylisoxazol-4-yl)-6-fluoro-N′-hydroxy-1-(4-hydroxyphenyl)-1H-indole-3-carboximidamide.

In some embodiments, the ERβ ligand is a compound selected from the ERβligands disclosed in U.S. Pat. No. 8,334,280, hereby incorporated byreference.

In some embodiments, the ERβ ligand is2-(3-fluoro-4-hydroxyphenyl)-7-vinyl-1,3-benzoxazol-5-ol (ERB-041;Wyeth). The ERβ ligand may be a substituted benzoxazole, such as any ofthe compounds disclosed in U.S. Pat. No. 6,794,403 or U.S. PatentApplication Publication No. 2011/0212923 (each of which is herebyincorporated by reference).

The ERβ ligand may be 2-(5-hydroxy-1,3-benzoxazol-2-yl)benzene-1,4-diol; 3-(5-hydroxy-1,3-benzoxazol-2-yl)benzene-1,2-diol;2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol;2-(3-chloro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol;2-(2-chloro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol;2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-6-ol;2-(3-tert-butyl-4-hydroxyphenyl)-1,3-benzoxazol-6-ol;2-(6-hydroxy-1,3-benzoxazol-2-yl)benzene-1,4-diol;3-(6-hydroxy-1,3-benzoxazol-2-yl)benzene-1,2-diol;4-(6-hydroxy-1,3-benzoxazol-2-yl)benzene-1,2-diol;2-(3-chloro-4-hydroxyphenyl)-1,3-benzoxazol-6-ol;4-(5-hydroxy-1,3-benzoxazol-2-yl)benzene-1,3-diol;4-(6-hydroxy-1,3-benzoxazol-2-yl)benzene-1,3-diol;6-chloro-2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol;6-bromo-2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol;6-chloro-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol;5-chloro-2-(4-hydroxyphenyl)-1,3-benzoxazol-6-ol;7-bromo-2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol;7-bromo-2-(2-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol;7-bromo-2-(2,3-difluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol;2-(4-hydroxyphenyl)-7-vinyl-1,3-benzoxazol-5-ol;7-(1,2-dibromoethyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol;7-(1-bromovinyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol;7-ethynyl-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol;2-(4-hydroxyphenyl)-7-propyl-1,3-benzoxazol-5-ol;7-butyl-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol;7-cyclopentyl-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol; ethyl5-hydroxy-2-(4-hydroxyphenyl)-1,3-benzoxazole-7-carboxylate:2-(4-hydroxyphenyl)-7-phenyl-1,3-benzoxazol-5-ol;2-(4-hydroxyphenyl)-7-methoxy-1,3-benzoxazol-5-ol;7-ethyl-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol;7-ethyl-2-(2-ethyl-4-hydroxyphenyl)-1,3-benzoxazol-5-ol;5-hydroxy-2-(4-hydroxyphenyl)-1,3-benzoxazole-7-carbaldehyde;7-(hydroxymethyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol;7-(bromomethyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol;[5-hydroxy-2-(4-hydroxy phenyl)-1,3-benzoxazol-7-yl]acetonitrile;7-(1-hydroxy-1-methylethyl)-2-(4-hydroxyphenyl)-[1,3-benzoxazol-5-ol];2-(4-hydroxyphenyl)-7-isopropenyl-1,3-benzoxazol-5-ol;2-(4-hydroxyphenyl)-7-isopropyl-[1,3-benzoxazol-5-ol];7-bromo-2-(4-hydroxy-3-(trifluoromethyl)phenyl)-1,3-benzoxazol-5-ol;7-(2-furyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol;2-(3-fluoro-4-hydroxyphenyl)-7-(2-furyl)-1,3-benzoxazol-5-ol;2-(4-hydroxyphenyl)-7-thien-2-yl-1,3-benzoxazol-5-ol;2-(4-hydroxyphenyl)-7-(1,3-thiazol-2-yl)-1,3-benzoxazol-5-ol;2-(3-fluoro-4-hydroxyphenyl)-5-hydroxy-1,3-benzoxazole-7-carbonitrile;4-bromo-2-(4-hydroxyphenyl)-7-methoxy-1,3-benzoxazol-5-ol;4,6-bisbromo-2-(4-hydroxy phenyl)-7-methoxy-1,3-benzoxazol-5-ol; or7-bromo-2-(3,5-difluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol.

The ERβ ligand may be 2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol,2-(3-chloro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol,2-(3-fluoro-4-hydroxyphenyl)-7-vinyl-1,3-benzoxazol-5-ol,2-(2-chloro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol,2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-6-ol,2-(3-tert-butyl-4-hydroxyphenyl)-1,3-benzoxazol-6-ol,2-(3-chloro-4-hydroxyphenyl)-1,3-benzoxazol-6-ol,6-chloro-2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol,6-bromo-2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol,6-chloro-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,5-chloro-2-(4-hydroxyphenyl)-1,3-benzoxazol-6-ol,7-bromo-2-(3-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol,7-bromo-2-(2-fluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol,7-bromo-2-(2,3-difluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol,2-(4-hydroxyphenyl)-7-vinyl-1,3-benzoxazol-5-ol,7-(1,2-dibromoethyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,7-(l-bromovinyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,7-ethynyl-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,2-(4-hydroxyphenyl)-7-propyl-1,3-benzoxazol-5-ol,7-butyl-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,7-cyclopentyl-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol, ethyl5-hydroxy-2-(4-hydroxyphenyl)-1,3-benzoxazole-7-carboxylate,2-(4-hydroxyphenyl)-7-phenyl-1,3-benzoxazol-5-ol,2-(4-hydroxyphenyl)-7-methoxy-1,3-benzoxazol-5-ol,7-ethyl-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,7-ethyl-2-(2-ethyl-4-hydroxyphenyl)-1,3-benzoxazol-5-ol,5-hydroxy-2-(4-hydroxyphenyl)-1,3-benzoxazole-7-carbaldehyde,7-(hydroxymethyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,7-(bromomethyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,[5-hydroxy-2-(4-hydroxyphenyl)-1,3-benzoxazol-7-yl] acetonitrile,7-(1-hydroxy-1-methylethyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,2-(4-hydroxyphenyl)-7-isopropenyl-1,3-benzoxazol-5-ol,2-(4-hydroxyphenyl)-7-isopropyl-1,3-benzoxazol-5-ol,7-bromo-2-(4-hydroxy-3-(trifluoromethyl)phenyl)-1,3-benzoxazol-5-ol,7-(2-furyl)-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol,2-(3-fluoro-4-hydroxyphenyl)-7-(2-furyl)-1,3-benzoxazol-5-ol,2-(4-hydroxyphenyl)-7-thien-2-yl-1,3-benzoxazol-5-ol,2-(4-hydroxyphenyl)-7-(1,3-thiazol-2-yl)-1,3-benzoxazol-5-ol,2-(3-fluoro-4-hydroxyphenyl)-5-hydroxy-1,3-benzoxazole-7-carbonitrile,4-bromo-2-(4-hydroxyphenyl)-7-methoxy-1,3-benzoxazol-5-ol,4,6-dibromo-2-(4-hydroxyphenyl)-7-methoxy-1,3-benzoxazol-5-ol, or7-bromo-2-(3,5-difluoro-4-hydroxyphenyl)-1,3-benzoxazol-5-ol.

In some embodiments, the ERβ ligand is a compound selected from the ERβligands disclosed in U.S. Patent Application Publication No.2007/0021495 or 2013/0274344, each of which is hereby incorporated byreference.

In some embodiments, the ERβ ligand is a halogen-substitutedphenyl-2H-indazole, such as indazole chloride (see, e.g., Moore, S. M.et al. Proc. Nat'l Acad. Sci. USA 111(5):18061-66 (2014), herebyincorporated by reference).

In some embodiments, the ERβ ligand is not estriol. The ERβ ligand maybe a non-steroidal compound. In some embodiments, the ERβ ligand is nota steroid hormone.

In certain aspects, the invention relates to methods for reducing thenumber or severity of at least one clinical sign or symptom of aneurodegenarative disease in a patient, comprising administering to thepatient an estrogen receptor beta ligand. e.g., wherein the estrogenreceptor beta ligand is a compound having the structure of formula I.

In certain embodiments, the invention relates to methods for preservingneurons in a patient afflicted with a neurodegenerative disease,comprising administering to the patient an estrogen receptor betaligand, e.g., wherein the estrogen receptor beta ligand is a compoundhaving the structure of formula I. Preserving neurons may comprisepreserving spinal cord axons.

In certain embodiments, the invention relates to methods for preservingmyelinating oligodendrocytes in a patient afflicted with aneurodegenarative disease, comprising administering to the patient anestrogen receptor beta ligand, e.g., wherein the estrogen receptor betaligand is a compound having the structure of formula I.

In certain embodiments, the invention relates to methods for preservingaxon myclination in a patient afflicted with a neurodegenerativedisease, comprising administering to the patient an estrogen receptorbeta ligand, e.g., wherein the estrogen receptor beta ligand is acompound having the structure of formula I.

In certain embodiments, the invention relates to methods for stimulatingaxon remyelination in a patient afflicted with a neurodegenerativedisease, comprising administering to the patient an estrogen receptorbeta ligand, e.g., wherein the estrogen receptor beta ligand is acompound having the structure of formula I.

In certain embodiments, the invention relates to methods reducing axonalloss in a patient afflicted with a neurodegenerative disease, comprisingadministering to the patient an estrogen receptor beta ligand, e.g.,wherein the estrogen receptor beta ligand is a compound having thestructure of formula I.

In certain embodiments, the invention relates to methods for reducingsynaptic loss in a patient afflicted with a neurodegenerative disease,comprising administering to the patient an estrogen receptor betaligand, e.g., wherein the estrogen receptor beta ligand is a compoundhaving the structure of formula I.

In certain embodiments, the invention relates to methods for reducingneuronal cell loss in a patient afflicted with a neurodegenerativedisease, comprising administering to the patient an estrogen receptorbeta ligand, e.g., wherein the estrogen receptor beta ligand is acompound having the structure of formula I.

In certain embodiments, the invention relates to methods for reducingbrain volume loss in a patient afflicted with a neurodegenerativedisease, comprising administering to the patient an estrogen receptorbeta ligand, e.g., wherein the estrogen receptor beta ligand is acompound having the structure of formula I.

In certain embodiments, the invention relates to methods for reducingcerebral cortex volume loss in a patient afflicted with aneurodegenerative disease, comprising administering to the patient anestrogen receptor beta ligand, e.g., wherein the estrogen receptor betaligand is a compound having the structure of formula I.

In certain embodiments, the invention relates to methods for reducingcerebellum volume loss in a patient afflicted with a neurodegenerativedisease, comprising administering to the patient an estrogen receptorbeta ligand, e.g., wherein the estrogen receptor beta ligand is acompound having the structure of formula I.

In certain embodiments, the invention relates to methods for reducinglocal central nervous system inflammation in a patient afflicted with aneurodegenerative disease, comprising administering to the patient anestrogen receptor beta ligand, e.g., wherein the estrogen receptor betaligand is a compound having the structure of formula I. The reduction ininflammation may be a result of a reduced number of macrophages.

In certain embodiments, the invention relates to methods for treatingone or more symptoms of multiple sclerosis in a patient in need thereof,comprising administering to the patient an effective amount of anestrogen receptor beta ligand, e.g., wherein the estrogen receptor betaligand is a compound having the structure of formula I The one or moresymptoms may include dystaxia, ataxia, or another motor disability.

The compound may be administered at a dose of about 1-100 mg/kg/day. Theestrogen receptor beta ligand may be a compound having the structure offormula I, and it may be administered at a dose sufficient to achieve amean blood concentration of the compound between 1 ng/ml and 1000 ng/ml.For example, the compound may be administered at a dose sufficient toachieve a mean blood concentration of the compound between 10 ng/ml and50 ng/ml, between 25 ng/ml and 75 ng/ml, between 50 ng/ml and 100 ng/ml,between 75 ng/ml and 125 ng/ml, between 100 ng/ml and 150 ng/ml, between125 ng/ml and 175 ng/ml, between 100 ng/ml and 200 ng/ml, between 150ng/ml and 250 ng/ml, between 200 ng/ml and 300 ng/ml, between 250 ng/mland 350 ng/ml, between 300 ng/ml and 400 ng/ml, between 350 ng/ml and450 ng/ml, between 400 ng/ml and 500 ng/ml, between 450 ng/ml and 650ng/ml, between 550 ng/ml and 750 ng/ml, between 650 ng/ml and 850 ng/ml,between 750 ng/ml and 950 ng/ml, or between 850 ng/ml and 1050 ng/ml. Insome embodiments, the compound may be administered at a dose sufficientto achieve a mean blood concentration of the compound between 100 ng/mland 200 ng/ml. In other embodiments, the compound may be administered ata dose sufficient to achieve a mean blood concentration of the compoundbetween 10 ng/ml and 20 ng/ml. In some embodiments, the compound isadministered at a dose sufficient to achieve a mean blood concentrationof the compound between 10 ng/ml and 500 ng/ml. The compound may beadministered at a dose sufficient to achieve a mean blood concentrationof the compound between 100 ng/ml and 200 ng/ml.

In certain embodiments, the ligand is administered on a daily basis.

The neurodegenerative disease may be Alzheimer's disease, Parkinson'sdisease, multiple sclerosis, stroke, amyotrophic lateral sclerosis,cerebellar ataxia, frontotemporal dementia, prion disease. Huntington'sdisease, cerebral ischemia, cerebral dementia syndrome,infection-induced neurodegeneration disorders (e.g.,AIDS-encephalopathy, Creutzfeld-Jakob disease, encephalopathics inducedby rubiola and herpes viruses and borrelioses), metabolic-toxicneurodegenerative disorders (such as hepatic-, alcoholic-, hypoxic-,hypo-, or hyperglycemicaly-induced encephalopathies), encephalopathiesinduced by solvents or pharmaceuticals, trauma-induced brain damage, orspinal cord injury.

In some embodiments, the neurodegenerative disease is multiple sclerosisor clinically isolated syndrome. The neurodegenerative disease may bemultiple sclerosis, and the patient may have one or more ofrelapsing-remitting multiple sclerosis, secondary-progressive multiplesclerosis, primary-progressive multiple sclerosis andprogressive-relapsing multiple sclerosis.

The method may further comprise administering to the patient animmunotherapeutic agent. The immunotherapeutic agent may initiated atthe same time or about the same time as initiation of treatment with theestrogen receptor beta ligand. Alternatively, the treatment with theestrogen receptor beta ligand may be initiated after the initiation oftreatment with the immunotherapeutic agent.

The immunotherapeutic agent may be selected from interferon-beta 1a,interferon-beta 1b, pegylated interferon-beta-1a, glatiramer acetate,natalizumab, mitoxantrone, fingolimod, teriflunomide, dimethyl fumarate,mycophenolate mofetil, pacitaxel, cyclosporine, corticosteroids,azathioprine, cyclophosphamide, methotrexate, cladribine,4-aminopyridine, and tizanidine. In some embodiments, theimmunotherapeutic agent is selected from interferon-beta 1a,interferon-beta 1b, pegylated interferon-beta-1a, glatiramer acetate,natalizumab, mitoxantrone, fingolimod, teriflunomide, and dimethylfumarate.

In some embodiments, the amount of the immunotherapeutic agentadministered in combination with the estrogen receptor beta ligand isless than a therapeutically effective amount when the immunotherapeuticagent is administered alone (e.g., 20 mg glatiramer five times a weekinstead of 40 mg three times a week or 20 mg daily; 0.4 mg fingolimoddaily instead of 0.5 mg daily; 200 mg dimethyl fumarate daily instead of240 mg daily; or 0.25 mg interferon beta-1b every third day instead of0.25 mg every other day). In some embodiments, the dose for glatirameracetate (Copaxone®) is 20 mg s.c. six times a week, five times a week,four times a week, or three times a week. In some embodiments, the dosefor fingolimod (Gilenya®) is 0.45 mg p.o. daily, 0.40 mg p.o. daily,0.35 mg p.o. daily, 0.30 mg p.o. daily, or 0.25 mg p.o. daily. In someembodiments, the dose for dimethyl fumarate (Tecfidera®) is 220 mg p.o.daily, 200 mg p.o. daily, 180 mg p.o. daily, 160 mg p.o. daily, 150 mgp.o. daily, 140 mg p.o. daily, or 120 mg p.o. daily. In someembodiments, the dose for interferon beta-1a (Avonex@) is 27, 26, 25,24, 23, 22, 21, 20, 19, 18, 17, 16, or 15 μg i.m. weekly, or 30 μgintramuscularly (i.m.) every 8, 9, 10 or 11 days. In some embodiments,the dose for interferon beta-1a (Rebif®) is 40, 39, 38, 37, 36, 35, 34,33, 3231, 30, 29, 28, 27, 26, 25, 24, 23, or 22 μg s.c. three days aweek, or 44 g s.c. two days a week. In some embodiments, the dose forinterferon beta-1b (Betaseron® or Extavia®) is 0.225, 0.200, 0.180,0.175, 0.170, 0.160, 0.150, 0.140, 0.130, or 0.125 mg s.c. every otherday, or 0.25 mg s.c. every third day.

In some embodiments, the patient is not being treated with anotherimmunotherapeutic agent (besides the estrogen receptor beta ligand). Theestrogen receptor beta ligand of formula I (or compounds of similarstructure and function) may be used as a monotherapy in latersecondary-progressive multiple sclerosis and primary-progressivemultiple sclerosis, in which the inflammatory component of the diseaseis small. Additionally, the estrogen receptor beta ligand of formula I(or compounds of similar structure and function) may be used as amonotherapy (e.g., without conjoint administration of ananti-inflammatory therapy) for treating relapsing-remitting multiplesclerosis and early secondary-progressive multiple sclerosis because theestrogen receptor beta ligand of formula I has both neuroprotective andanti-inflammatory properties.

The patient may be a rodent or primate. In some embodiments, the patientis a human.

The patient may be male or female. In some embodiments, the patient isfemale; in other embodiments, male.

In certain aspects, the invention relates to methods of treatingmultiple sclerosis in a subject having a loss of brain gray matter ofgreater than about 0.3% in a period of at least about 6 months,comprising administering an estrogen receptor beta ligand to the subjectdaily, wherein: the estrogen receptor beta ligand is a compound havingthe structure of formula; and the compound is administered at a dosesufficient to achieve a mean blood concentration of the compound between10 ng/ml and 500 ng/ml. The loss of brain gray matter may be measured byMRI In some embodiments, the treatment slows or eliminates a loss ofbrain gray matter. The loss of gray matter may be located in one or moreof total brain, cerebral cortex, cerebellum, thalamus, caudate nucleus,and putamen.

In some embodiments, the invention relates to methods of treating aneurodegenerative disease, comprising administering to a subject in needthereof a first treatment regimen; assessing a change in total braingray matter volume of the subject over a period of time; and changingthe treatment regimen if the total brain gray matter volume decreases byat least about 0.3 percent between a first assessment and a secondassessment about 6 months after the first assessment, by at least about0.6 percent between a first assessment and a second assessment about oneyear after the first assessment, or by at least about 1.0 percentbetween a first assessment and a second assessment about two years afterthe first assessment. Changing the treatment regimen may compriseadministering an effective amount of an estrogen receptor beta ligand tothe subject in addition to the first treatment regimen.

In some embodiments, the invention relates to methods of treating aneurodegenarative disease, comprising administering to a subject in needthereof a first treatment regimen; assessing a change in gray mattervolume of at least two brain regions of the subject over a period oftime: and changing the treatment regimen if the gray matter volume ofthe at least two brain regions decreases by at least about 0.3 percentbetween a first assessment and a second assessment about 6 months afterthe first assessment, by at least about 0.6 percent between a firstassessment and a second assessment about one year after the firstassessment, or by at least about 1.0 percent between a first assessmentand a second assessment about two years after the first assessment.Changing the treatment regimen may comprise administering an effectiveamount of an estrogen receptor beta ligand to the subject in addition tothe first treatment regimen. The at least two brain regions may beselected from cerebral cortex, cerebellum, thalamus, caudate nucleus,putamen, and any combination thereof. In some embodiments, the at leastone brain region is cerebral cortex.

Assessing may comprise performing brain magnetic resonance imaging(MRI). The first assessment may be performed before, at the same timeas, or at about the same time as initiating the treatment regimen. Thefirst assessment may be performed after the treatment regimen isinitiated. The second assessment may be performed at least about sixmonths, at least about one year, at least about 18 months, or at leastabout two years after said first assessment. In some embodiments, thesecond assessment is performed about one year after said firstassessment.

The neurodegenerative disease may be Alzheimer's disease, Parkinson'sdisease, multiple sclerosis, stroke, amyotrophic lateral sclerosis,cerebellar ataxia, frontotemporal dementia, prion disease, Huntington'sdisease, cerebral ischemia, cerebral dementia syndrome,infection-induced neurodegeneration disorders (e.g.,AIDS-encephalopathy, Creutzfeld-Jakob disease, encephalopathies inducedby rubiola and herpes viruses and borrelioses), metabolic-toxicneurodegenerative disorders (such as hepatic-, alcoholic-, hypoxi-,hypo-, or hyperglycemically-induced encephalopathies), encephalopathicsinduced by solvents or pharmaceuticals, trauma-induced brain damage, orspinal cord injury.

In some embodiments, the neurodegenerative disease is multiple sclerosisor clinically isolated syndrome. The subject may have one or more ofrelapsing-remitting multiple sclerosis, secondary-progressive multiplesclerosis, primary-progressive multiple sclerosis andprogressive-relapsing multiple sclerosis.

In some embodiments, the treatment regimen comprises administering tothe subject an immunotherapeutic agent. The immunotherapeutic agent maybe selected from interferon-beta 1a, interferon-beta 1b, pegylatedinterferon-beta-1a, glatiramer acetate, natalizumab, mitoxantrone,fingolimod, teriflunomide, dimethyl fumarate, mycophenolate mofetil,paclitaxel, cyclosporine, corticosteroids, azathioprine,cyclophosphamide, methotrexate, cladribine, 4-aminopyridine, andtizanidine. In some embodiments, the immunotherapeutic agent is selectedfrom interferon-beta 1a, interferon-beta 1b, pegylatedinterferon-beta-1a, glatiramer acetate, natalizumab, mitoxantrone,fingolimod, teriflunomide, and dimethyl fumarate. In some embodiments,the amount of the immunotherapeutic agent administered in combinationwith the estrogen receptor beta ligand is less than a therapeuticallyeffective amount when the immunotherapeutic agent is administered alone(e.g., 20 mg glatiramer five times a week instead of 40 mg three times aweek or 20 mg daily; 0.4 mg fingolimod daily instead of 0.5 mg daily;200 mg dimethyl fumarate daily instead of 240 mg daily; or 0.25 mginterferon beta-1b every third day instead of 0.25 mg every other day).In some embodiments, the dose for glatiramer acetate (Copaxone®) is 20mg s.c. six times a week, five times a week, four times a week, or threetimes a week. In some embodiments, the dose for fingolimod (Gilenya®) is0.45 mg p.o. daily, 0.40 mg p.o. daily, 0.35 mg p.o. daily, 0.30 mg p.o.daily, or 0.25 mg p.o. daily. In some embodiments, the dose for dimethylfumarate (Tecfidera®) is 220 mg p.o. daily, 200 mg p.o. daily, 180 mgp.o. daily, 160 mg p.o. daily, 150 mg p.o. daily, 140 mg p.o. daily, or120 mg p.o. daily. In some embodiments, the dose for interferon beta-1a(Avonex®) is 27, 26, 25, 24, 23, 22, 21, 20, 19, 18, 17, 16, or 15 gi.m. weekly, or 30 μg intramuscularly (i.m.) every 8, 9, 10 or 11 days.In some embodiments, the dose for interferon beta-1a (Rebif®) is 40, 39,38, 37, 36, 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, 24, 23, or 22 μgs.c. three days a week, or 44 μg s.c. two days a week. In someembodiments, the dose for interferon beta-1b (Betaseron® or Extavia®) is0.225, 0.200, 0.180, 0.175, 0.170, 0.160, 0.150, 0.140, 0.130, or 0.125mg s.c. every other day, or 0.25 mg s.c. every third day.

In certain embodiments, the subject is receiving an immunotherapeuticagent and has cognitive disability. For example, if a subject scoresbelow 50 on PASAT, and optionally if such low score is verified uponretest within about one week to one month, then the subject may bedeemed to have cognitive disability. In accordance with the invention,this cognitive disability is treated with an estrogen receptor betaligand, and, in certain embodiments, followed up with further reteste.g., about six months from the start of an estrogen receptor betaligand treatment, such as to achieve an increase in test score of atleast 3 points.

In certain embodiments, the subject is receiving an immunotherapeuticagent and has progressive walking disability. For example, the subjectperforms a 25 foot walk test, e.g., at 0 months (baseline), 6 months, 1year, and/or 2 years. If there is documented worsening in walking (takesmore seconds e.g., by 20 percent as compared to baseline, and thisworsening is confirmed on a repeated walk test, e.g., about 3 monthslater, then the subject is deemed to have progressive worsening inwalking. In accordance with the invention, this progressive walkingdisability is treated with an estrogen receptor beta ligand, and, incertain embodiments, followed up with repeat walking test, e.g., atabout 1 year or 2 years from the start of an estrogen receptor betaligand treatment, such as to stabilize or halt any further worsening inwalking times.

In some embodiments, the first treatment regimen does not comprise anestrogen receptor beta ligand. The estrogen receptor beta ligand may bea compound having the structure of formula I. The compound of formula Iis called AC-186. In some embodiments the estrogen receptor beta ligandis a compound that is substantially similar in structure and function toAC-186, such as one of the compounds disclosed in PCT Patent PublicationWO 2013/017619 A1 (incorporated herein by reference).

In some embodiments, the estrogen receptor beta ligand is a compoundselected from the compounds disclosed in U.S. Patent ApplicationPublications 2012/0202861 A1 or 2013/0 131061 A1 (incorporated herein byreference). In some embodiments, the estrogen receptor beta ligand isKBRV1 or KBRV2 (Karo Bio. Huddinge, Sweden).

The wherein estrogen receptor beta ligand may be a compound having thestructure of formula I, and it may be administered at a dose sufficientto achieve a mean blood concentration of the compound between 1 ng/mland 1000 ng/ml. In some embodiments, the compound is administered at adose sufficient to achieve a mean blood concentration of the compoundbetween 10 ng/ml and 500 ng/ml. The compound may be administered at adose sufficient to achieve a mean blood concentration of the compoundbetween 100 ng/ml and 200 ng/ml.

In certain embodiments, the ligand is administered on a daily basis.

The subject may be a rodent or primate. In some embodiments, the subjectis a human.

The subject may be male or female. In some embodiments, the subject isfemale.

This above description and following examples are not to be taken in alimiting sense, but are made merely for the purpose of illustrating thegeneral principles of the invention. The section titles and overallorganization of the present detailed description are for the purpose ofconvenience only and are not intended to limit the present invention.

Example 1 Materials and Methods.

Animals. C57BL6 and NOD mice 8 weeks old were purchased from JacksonLaboratories (Bar Harbor, Me.). Animals were maintained underenvironmentally controlled conditions in a 12-hour light/dark cycle withaccess to food and water ad libitum. All procedures were done inaccordance with the guidelines of the National Institutes of Health andthe Chancellor's Animal Research Committee of the University ofCalifornia, Los Angeles Office for the Protection of Research Subjects.

Reagents. AC-186 was dissolved in either Miglyol 812N liquid oil (SasolNorth America) or sesame oil (Sigma Aldrich) as following concentration;1.5 mg/mL for 3 mg/kg group, 5 mg/ml for 10 mg/kg group, and 15 mg/mlfor 30 mg/kg group. Diarylpropionitrile (DPN, Tocris Biosciences) wasdissolved in 10% molecular-grade ethanol and diluted with 90% of eitherMiglyol 812N liquid oil or sesame oil.

EAE and Treatments. Animals were injected subcutaneously with MyelinOligodendrocyte Glycoprotein (MOG), amino acids 35-55 (200 g/animal,American Peptides), emulsified in complete Freund's adjuvant (CFA) andsupplemented with Mycobacterium Tuberculosis H37ra (300 μg/animal, DifcoLaboratories), over two sites drained by let inguinal and auxiliarylymph nodes in a total volume of 0.1 ml/mouse. One week later, a boosterimmunization was delivered over contra lateral lymph nodes. Pertussistoxin (500 ng/mouse) (List Biological Laboratories, Inc.) was injectedintraperitoneally on days 0 and 2. Animals were monitored daily for EAEsigns based on a standard EAE 0-5 scale scoring system: 0—healthy,1—complete loss of tail tonicity, 2—loss of righting reflex, 3—partialparalysis, 4—complete paralysis of one or both hind limbs, and5—moribund. Animals received 0.05 ml of either of 1.5, 5, 15 mg/mLAC-186, vehicle (sesame oil or Miglyol 812N liquid oil), or DPN (8mg/kg/day) via subcutaneous injections every other day. Animals weretreated with AC-186 or vehicle after EAE induction. Specifically,treatment was initiated at the first clear signs of clinical disease(EAE day 13-15), and continued to the endpoint of the experiment.

Rotarod Testing. Motor behavior was tested up to two times per week foreach mouse using a rotarod apparatus (Med Associates Inc., St. Albans,Vt.). Briefly, animals were placed on a rotating horizontal cylinder fora maximum of 200 seconds. The amount of time the mouse remained walkingon the cylinder, without falling, was recorded. Each mouse was tested ona speed of 3-30 rpm and given three trials for any given day. The threetrials were averaged to report a single value for an individual mouse,and then averages were calculated for all animals within a giventreatment group. The first two trial days prior to immunization (day 0),served as practice trials.

Histological Preparation. Mice were exposed to a lethal dose ofisoflurane and perfused transcardially with ice-cold 1×PBS for 8-15 min,followed by 10% formalin for 10-15 min. Spinal cords and brains weredissected and submerged in 10% formalin overnight at 4° C., followed by30% sucrose in PBS for 24 h. Tissues were embedded in 75% gelatin/15%sucrose solution for cryostat sectioning then post-fixed overnight in10% formalin and cryoprotected in 30% sucrose. The embedded tissues werestored in −80° C. after flash frozen in dry ice. 40 μm thickfree-floating spinal cord cross-sections, and sagittal brain sectionswere obtained with a microtome b cryostat (model IM505E) at −20° C.Tissues were collected serially and stored in 1×PBS with 1% sodium azidein 4° C. until immunohistochemistry.

Immunofluorescence. Prior to histological staining, 40-mm thickfree-floating sections were thoroughly washed with IX PBS to removeresidual sodium azide. In the case of anti-MBP labeling, tissue sectionswere processed with an additional 1 hour incubation with 5% glacialacetic acid in 100-proof ethanol at room temperature (RT). After washingtissue sections were permeabilized with 0.3% TritonX-100 and 2% normalgoat scrum in 1×PBS for 30 minutes at room temperature and blocked with10% normal goat serum in 1×PBS for 1 hour. Tissues were then incubatedwith primary antibodies overnight in 4° C. The following primaryantibody (Ab) were used: Rat anti-MBP (Millipore) at 1:1000 dilutions,Rabbit anti-NF200 (Sigma Aldrich) at 1:750 dilutions, Rabbitanti-beta-APP (Life Technologies) at 1:200 dilutions, Mouse anti-NeuN at1:1000 dilutions (Millipore), Rabbit anti-PSD95, and Rabbitanti-Synapsin 1 at 1:500 dilutions (Millipore), and Rat anti-CD45 at1:1500 dilutions (Millipore). The next day tissues were washed andincubated with secondary antibodies conjugated to Cy5 or Cy3 (Millipore)for 1 hour at room temperature. A nuclear stain DA PI (2 ng/mL;Molecular Probes) was added 10 minutes prior to final washes aftersecondary Ab incubation. Sections were mounted on slides, allowed tosemi-dry, and cover slipped in fluoromount G (Fisher Scientific).IgG-control experiments were performed for all primary Ab, and onlynon-immunoreactive tissues under these conditions were analyzed.

Chromagen Immunohistochemistry. Tissue sections were thoroughly washedwith 1×PBS to remove residual sodium azide and treated with 3% hydrogenperoxide for 30 minutes at room temperature and then simultaneouslyblocked with 10% NGS and permeabilized with 0.3% Triton X-100 in 1×PBSfor 1 hour at room temperature. Tissues were then incubated with primaryantibodies overnight in 4° C. The following primary antibodies wereused: Rat anti-CD3 at 1:2000 dilutions (BD Pharmigen), anti-CalbindinD28K at 1:1000 dilutions (Millipore), and Rabbit anti-Iba1 at 1:10000dilutions (Wako Chemicals), were added for 2 hour at room temperature,and then placed in 4° C. overnight. Tissue sections then followed withsecondary Ab labeling at 1:1000 dilutions (Vector labs) for 1 hour atroom temperature and then with Avidin-Biotin Conjugation solution(Vector Labs) for 1 hour at room temperature. Tissue sections weretreated with DAB peroxidase substrate (Vector labs) according tomanufacturer instructions. IgG-control experiments were performed forall primary Ab. and only non-immunoreactive tissues under theseconditions were analyzed.

Microscopy and Quantification. Three spinal cord cross-sections at theC5-12 level from each mouse (n=3-6) were captured under microscope at10× magnification. To quantify demyelination in the spinal cord andcerebellum, white matter was manually delineated on the basis of DAP1staining, and MBP staining intensity was calculated and reported in thesampled area. Axonal damage was assessed by counting beta-APP cells in aconfocal 10× microscope in spinal cord. Axonal densities were calculatedby counting the number of NF200⁺ or NeuN⁺ neuronal cells in a Oxconfocal image in the sampled area of the captured tissue section.Cerebellar Purkinje (Calbindin⁺) cells were manually counted using abrightfield 10× microscope over the entire sagittal cerebellum. PSD-95and Synapsin1 density was measured and reported as a percentage of thesampled area. CD45⁺ CD3⁺, and Iba1⁺ cells in spinal cord cross-sectionswere manually quantified under either of a confocal 10× microscope forCD45′ cells or a brightfield 10× microscope for CD3′ and Iba1⁺ cells.Histological Preparation. Mice were exposed to a lethal dose ofisoflurane and perfused transcardially with ice-cold 1′×PBS for 8-15min, followed by 10% formalin for 10-15 min. Spinal cords and brainswere dissected and submerged in 10% formalin overnight at 4° C.,followed by 30% sucrose in PBS for 24 h. Tissues were embedded in 75%gelatin/15% sucrose solution for cryostat sectioning then post-fixedovernight in 10% formalin and cryoprotected in 30% sucrose. The embeddedtissues were stored in −80° C. after flash frozen in dry ice. 40 μmthick free-floating spinal cord cross-sections, and sagittal brainsections were obtained with a microtome b cryostat (model HM505E) at−20° C. Tissues were collected serially and stored in 1×PBS with 1%sodium azide in 4° C. until immunohistochemistry.

Immunofluorescence. Prior to histological staining, 40-mm thickfree-floating sections were thoroughly washed with 1×PBS to removeresidual sodium azide. In the case of anti-MBP labeling, tissue sectionswere processed with an additional 1 hour incubation with 5% glacialacetic acid in 100-proof ethanol at room temperature (RT). After washingtissue sections were permeabilized with 0.3% TritonX-100 and 2% normalgoat serum in 1×PBS for 30 minutes at room temperature and blocked with10% normal goat serum in 1×PBS for 1 hour. Tissues were then incubatedwith primary antibodies overnight in 4° C. The following primaryantibody (Ab) were used: Rat anti-MBP (Millipore) at 1:1000 dilutions.Rabbit anti-NF200 (Sigma Aldrich) at 1:750 dilutions, Rabbitanti-beta-APP (Life Technologies) at 1:200 dilutions, Mouse anti-NeuN at1:1000 dilutions (Millipore). Rabbit anti-PSD95, and Rabbitanti-Synapsin1 at 1:500 dilutions (Millipore), and Rat anti-CD45 at1:1500 dilutions (Millipore). The next day tissues were washed andincubated with secondary antibodies conjugated to Cy5 or Cy3 (Millipore)for 1 hour at room temperature. A nuclear stain DAP1 (2 ng/mL; MolecularProbes) was added 10 minutes prior to final washes after secondary Abincubation. Sections were mounted on slides, allowed to semi-dry, andcover slipped in fluoromount G (Fisher Scientific). IgG-controlexperiments were performed for all primary Ab, and onlynon-immunoreactive tissues under these conditions were analyzed.

Chromagen Immunochemistry. Tissue sections were thoroughly washed with1×PBS to remove residual sodium azide and treated with 3% hydrogenperoxide for 30 minutes at room temperature and then simultaneouslyblocked with 10% NGS and permeabilized with 0.3% Triton X-100 in 1×PBSfor 1 hour at room temperature. Tissues were then incubated with primaryantibodies overnight in 4° C. The following primary antibodies wereused: Rat anti-CD3 at 1:2000 dilutions (BD Pharmigen), anti-CalbindinD28K at 1:1000 dilutions (Millipore), and Rabbit anti-Iba1 at 1:10000dilutions (Wako Chemicals), were added for 2 hour at room temperature,and then placed in 4′C overnight. Tissue sections then followed withsecondary Ab labeling at 1:1000 dilutions (Vector labs) for 1 hour atroom temperature and then with Avidin-Biotin Conjugation solution(Vector Labs) for 1 hour at room temperature. Tissue sections weretreated with DAB peroxidase substrate (Vector labs) according tomanufacturer instructions. IgG-control experiments were performed forall primary Ab, and only non-immunoreactive tissues under theseconditions were analyzed.

Microscopy and Quantification. Three spinal cord cross-sections at theC5-12 level from each mouse (n=3-6) were captured under microscope at 1×magnification. To quantify demyelination in the spinal cord andcerebellum, white matter was manually delineated on the basis of DAPIstaining, and MBP staining intensity was calculated and reported in thesampled area. Axonal damage was assessed by counting beta-APP⁺ cells ina confocal 10× microscope in spinal cord. Axonal densities werecalculated by counting the number of NF200⁻ or NeuN⁺ neuronal cells in a10× confocal image in the sampled area of the captured tissue section.Cerebellar Purkinje (Calbindin⁺) cells were manually counted using abrightfield 10× microscope over the entire sagittal cerebellum. PSD-95and Synapsin 1 density was measured and reported as a percentage of thesampled area. CD45⁺, CD3⁺, and Iba1⁺ cells in spinal cord cross-sectionswere manually quantified under either of a confocal 10× microscope forCD45⁺ cells or a brightfield 10× microscope for CD3⁺ and Iba1⁺ cells.

Example 2

Three doses of the AC-186 compound were tested in C57BL/6 female mice:low (3 mg/kg, every other day), medium (10 mg/kg, every other day), andhigh (30 mg/kg, every other day), each injected subcutaneously in asesame oil vehicle. The ERβ ligand diarylpropionitrile (DPN) was testedas a positive control because DPN can partially ameliorate EAE andbecause the treatment effects in EAE experiments can vary based on thelevel of disease severity within an experiment. Direct comparisonsbetween treatments were only made between treatment groups within asingle EAE experiment and not between treatment groups of different EAEexperiments. DPN, however, was difficult to dissolve in sesame oil, andthus, DPN was first dissolved in ethanol and added to sesame oil at aconcentration of 10% ethanol in sesame oil. The AC-186 compounddissolved in sesame oil after five minutes on a nutating shaker. Thus,FIG. 1 shows graphs in which the vehicle consisted of sesame oil forAC-86 (all doses) and vehicle, while the DPN vehicle was 10% ethanol insesame oil. The low (3 mg/kg) dose of AC-186 had no effect on the EAEscore, the medium dose (10 mg/kg) displayed a trend toward improvement,and the high (30 mg/kg) dose displayed a significant effect inameliorating standard EAE clinical scores (p=0.0299) (FIGS. 1 and 2).The difference in improvement between the 30 mg/kg AC-186 group and allother groups increased with time.

To assess whether a more novel EAE outcome might be affected as well bythe high (30 mg/kg) dose, rotarod was also performed, as shown in FIG.3. Rotarod performance is likely more aligned with coordination andcerebellar function than standard EAE scores, which reflect principallywalking and spinal cord pathology. No significant effect was observedfor mice receiving 30 mg/kg AC-186 in sesame oil relative to thosereceiving vehicle only: however, very late in disease, at the time whenER beta ligands are known to start working, the performance curvestrended toward divergence, with AC-186 treated mice trending towardimproved performance. Further, the rotarod test is insensitive indetecting differences when the vehicle group performs well, and thevehicle group performed well in this case, staying on the rotarod forapproximately 150 seconds. Additionally, during the final twotime-points, on days 48 and 50, the AC-86 group performed perfectlyduring the 200 second test, and thus, a significant effect may have beenmasked by the experimental design.

Example 3

DPN protects against EAE better when administered in a Miglyol vehiclerather than in a sesame oil vehicle, and thus, the effect of the choiceof vehicle on the efficacy of AC-186 was assessed. Accordingly, theAC-186 compound was assessed in C57BL/6 males using sesame oil andmiglyol as vehicles. Male mice receiving AC-186 administered at 30 mg/kgin miglyol performed significantly better than mice receiving miglyolalone as assessed by both EAE score (FIG. 5) and rotarod performance(FIG. 6). In contrast, male mice receiving AC-186 administered at 30mg/kg in sesame oil did not perform significantly better than micereceiving sesame oil alone as assessed by EAE score (FIG. 5) and rotarodperformance (FIG. 6). Notably, the AC-186 solution dissolved morerapidly in miglyol than sesame oil, and miglyol could dissolve AC-186 bymerely pipetting for 30 seconds. In contrast, AC-186 requiredmixing/nutating for 5 minutes to dissolve the compound in sesame oil.

The 30 mg/kg dose of AC-186 dissolved in sesame oil did not displayefficacy in male C57BL6 mice, in contrast with the results obtained forfemale C57BL6 mice as described in Example 2.

Example 4

Sesame oil has been shown previously to have some nonspecificimmunostimulatory effects, and thus is not most commonly used as avehicle in EAE. As shown in FIG. 7, the different vehicle type does notaffect EAE differentially when given without an ERβ ligand. Rather, thedifferent vehicle type likely affects the ability of a given ERβ ligandto protect in EAE, with Miglyol enabling better EAE protection thansesame oil when either DPN or AC-186 are administered.

Example 5

The effect of AC-186 was assessed in female NOD mice using a MOG-inducedEAE model. The efficacy of AC-186 was tested in miglyol and sesame oilvehicles. In these models, AC-186 improved EAE scores relative tovehicle only for mice receiving 10 mg/kg or 30 mg/kg in miglyol(p<0.001), and the 30 mg/cg group trended toward increased efficacyrelative to the 10 mg/kg group (FIG. 8). In comparison, the positivecontrol DPN, which had not previously been tested in the NOD EAE model,also significantly ameliorated EAE, appearing similar to the diseasereduction observed with the 30 mg/kg dose of AC-186. In contrast, AC-186in sesame oil did not ameliorate EAE scores, consistent with data abovein the C57BL/6 male EAE experiment where AC-186 in miglyol amelioratedEAE while AC-186 in sesame oil did not.

Rotarod testing had never before been done in the NOD EAE model, androtarod scores were surprisingly poor with most mice staying on therotarod for only 0-100 seconds and none staying on for 125-200 seconds.These results were surprising in part because the EAE walking scoreswere reasonably, although not dramatically, severe in the moderate rangeof 2-3. These results may suggest that EAE in the NOD modelpreferentially affects cerebellar or other balance related pathways ascompared to EAE in the C57BL/6 model. The intervention produced nosignificant improvement in rotarod scores (FIG. 9).

Example 6

The effect of AC-186 was assessed in male NOD mice using a MOG-inducedEAE model. The efficacy of AC-186 was tested in miglyol vehicle only.AC-186 improved EAE scores relative to vehicle only for mice receiving30 mg/kg in miglyol (p<0.0001) (FIG. 10). Mice treated with AC-186displayed no significant improvement in performance in the rotarodexperiment relative to mice treated with vehicle only (FIG. 11).

Example 7

Two doses of the AC-186 compound were tested in C57BL/6 female mice:medium (10 mg/kg) and high (30 mg/kg), each in a miglyol vehicle. Boththe medium dose (10 mg/kg) and high dose (30 mg/kg) displayedsignificant efficacy in ameliorating the standard EAE clinical scoresrelative to the miglyol vehicle alone (p=0.0001)(FIG. 12). Additionally,the high dose (30 mg/kg) displayed significant efficacy in the rotarodexperiment relative to the miglyol vehicle alone (p=0.0005) (FIG. 13).

TABLE 1 Summary of standard EAE and rotarod scores Mouse Results StrainSex AC186 Vehicle Standard EAE Score Rotarod Score C57BL/6 F 30 mg/kgmiglylol **** *** C57BL/6 M 30 mg/kg miglylol **** **** NOD F 30 mg/kgmiglylol **** n.i. NOD M 30 mg/kg miglylol **** n.i. C57BL/6 F 10 mg/kgmiglylol **** (less robust that 30 mg/kg) n.s C57BL/6 M 10 mg/kgmiglylol N/A N/A NOD F 10 mg/kg miglylol **** n.i. NOD M 10 mg/kgmiglylol n.s. n.i. C57BL/6 F 30 mg/kg sesame * n.s. C57BL/6 M 30 mg/kgsesame n.s. n.s. NOD F 30 mg/kg sesame n.s. n.i. NOD M 30 mg/kg sesameN/A N/A C57BL/6 F 10 mg/kg sesame n.s. n.s. C57BL/6 F  3 mg/kg sesamen.s. n.s. n.s. = not significant; N/A = not applicable, not done; *p <0.05, ***p = 0.0004, ****p < 0.0001 n.i. = not informative; NOD femalesand NOD males have very poor rotarod performance, and thus the rotarodexperiment is not sensitive in the NOD strain.

Example 8

Neuroprotective effects of AC-186 were observed by NF200 and beta-APPstaining, with each of the 30 mg/kg and the 10 mg/kg doses (FIG. 14).C57BL/6 female mice were treated with AC-186 at 30 mg/kg/every other dayin miglyol vehicle, 10 mg/kg/every other day in miglyol vehicle or withmiglyol vehicle alone then underwent immunohistochemistry for axonal andmyelin integrity using antibody staining for NF200, beta-APP and MBP.NF200 indicated axonal integrity with decreases indicating axonal loss,beta-APP also indicated axonal integrity with increases indicatingaxonal damage, and MBP staining indicated myelin integrity withdecreases indicating demyelination during EAE. AC-186 30 mg/kg/everyother day treatment in EAE significantly preserved axon numbers (NF200)and reduced axonal damage (beta-APP), with a trend for sparing myelin(MBP). These beneficial effects on axons in spinal cord are consistentwith protective effects on clinical scores as assessed by EAE standardscores and rotarod performance above.

Neuropathology of spinal cords focusing on immune cells is shown in FIG.15. Regarding the assessment of immune cells by neuropathology, incontrast to previous experiments with DPN, which showed no significanteffect on CD45 staining in the CNS, AC-186 treatment at both the 10mg/kg and the 30 mg/kg doses reduced CD45 staining. To determine whichimmune cell population was reduced, Iba-1 stained cells with globoidmorphology were used to assess macrophages and CD3 staining was used toassess T lymphocytes. AC-186 treatment at both the 30 mg/kg and the 10mg/kg dose each reduced Iba-1 globoid cell staining, while neitherAC-186 dose affected levels of CD3 staining.

These data are consistent with protective effects on clinical EAEscores. Effects on myelin staining were much less striking as comparedto effects on axons as has been previously observed. Surprisingly, incontrast to previous experiments with DPN, which showed no effect onCD45 or Iba-1 globoid cell staining, AC-186 treatment reduced CD45 andIba-1 globoid cell staining. However, neither dose of AC-186 affectedlevels of CD3 staining (FIG. 15). Whether the effect of AC-186 onreducing Iba-1 globoid cells was due to a reduction in macrophageinfiltration into the CNS or due to a reduction in the transition of CNSresident cells to the globoid morphology as a reaction to less axonaldamage during AC-186 treatment remains unknown. The observation that Tlymphocyte levels were unaffected by AC-186 treatment suggests that theadaptive immune response is not affected by treatment like the innateimmune response is. Taken together, these results suggest that ER betaligands differ in their effect on CD45 and other immune marker staining,which can have therapeutic and mechanistic implications for thetreatment of MS subtypes.

MRI Methods

MRI Acquisition. Mice were anesthetized with isofluorance and theirheads secured with bite and car bars. Respiration rate was monitored andthe mice were maintained at 37° C. using a circulating water pump. Invivo magnetic resonance imaging was performed using a 200 mm horizontalbore 7.0 T Bruker imaging spectrometer with a micro-imaging gradientinsert with a maximum gradient strength of 100 G/cm and 30 mm birdcageRF coil (Bruker Instruments, Billerica, Mass.). An actively decoupledquadrature surface coil array was used for signal reception and a 72-mmbirdcage coil was used for transmission. Images were acquired andreconstructed using ParaVision 5.1 software, Imaging parameters were asfollows: rapid-acquisition with relaxation enhancement (RARE) sequence,matrix dimensions=256×128×64; field of view 3.84 cm×1.92 cm×0.96 cm;repetition time (TR)=3500 ms; apparent time to echo (apparentTE)=32 ms;echo train length=16; total scan time=37 mins. Spatial resolution was150 μm³ per voxel.

MRI Analysis. Images were skull-stripped using the Brain SurfaceExtractor (BSE) and residual non-brain signal was removed by a singleoperator manually editing the masks using BrainSuite 11a and bias-fieldinhomogeneitics removed using the N3 correction. After inhomogeneitycorrection, a minimum deformation atlas (MDA) was produced. Images werespatially and intensity normalized to the MDA using a rigid-bodytransformation and an intensity rescaling cost function in Alignlinear(AIR). This process permits the comparison of images in a standard spacecorrecting for both gross positional and intensity differences, yetpreserving anatomically significant local changes. Following creation ofthis atlas, cerebral cortices and cerebella were manually labeled on theatlas. The labels were then warped onto the individual spatiallynormalized images to produce standardized estimates of gray mattervolumes in individual subjects. All automated image processing wasperformed using the LONI Pipeline Processing Environment on an8-processor core Mac Pro computer (Apple. Cupertino, Calif.).

Cerebral cortex and cerebellum labels were based on the Mouse AtlasProject 2003 mouse brain atlas. For clarity and consistency, thecerebral cortex label was bounded ventrally by the plane inferior to themost anterior point of the corpus callosum at midline. Importantly, thislabel contained the somatosensory regions (primary and secondary) andthe motor cortex (primary and secondary). Additional anatomicalinformation was obtained from the Franklin and Paxinos mouse brain atlas(Franklin and Paxinos, 2008).

Statistics. Global and regional brain volume changes in EA E mice andcontrol animals were compared with repeated measures ANOVAs using SPSS22 (IBM, Armonk, N.Y.). If Mauchly's test indicated that the assumptionof sphericity had been violated (p 0.05, then the degrees of freedomwere corrected using the Huynh-Feldt estimates of sphericity. Regressionanalysis and Welch's t-tests were performed in Excel 2011 (Microsoft,Redmond, Wash.). All results are presented as mean±standard deviation.

Example 9

Female C57BL/6 mice treated with AC-186 at 30 mg/kg underwent in vivo,longitudinal MRI scanning at day 0, 30, and 60 after EAE induction. Themice treated with AC-186 displayed beneficial effects for both standardEAE clinical scores and for rotarod times, similar to the effectsobserved in Example 7 (FIG. 16). Whole brain, cerebral cortex andcerebellar volumes were determined at each EAE time point in femaleC57Bl/6 mice that were treated with either AC-186 (AC-186) or vehicle(EAE), as well as in age- and sex-matched healthy control mice (NOR).

Mice underwent in vivo, longitudinal MRI scanning at day 0, 30, and 60after EAE induction. Whole brain, cerebral cortex and cerebellar volumeswere determined at each EAE time point in female C57Bl/6 mice that weretreated with either AC-186 (AC-186, 30 mg/kg/every other day) or vehicle(EAE) as well as in age- and sex-matched healthy control mice (NOR).

NORMAL vs. EAE. Whole brain volumes of both NOR (black lines) and EAEvehicle mice were plotted against the disease duration (starting withthe scans prior to disease induction) (FIG. 17A). In order to quantifythe significance of the decreases in whole brain volume observed inindividual animals, a repeated-measures ANOVA was performed to assessthe effect of time on whole brain volume. Brain volume remained stableover time in the NOR group, but showed a gradual decrease in the EAEgroup (time×group interaction p=1.1×10⁻⁷). The volume of the whole brainof mice sixty days after disease induction (d60) was 490 mm³ (3.2 mm³)in NOR mice and 478 mm³ (3.7 mm³) in EAE mice, indicating a 4.7%decrease (p=5.3×10⁻⁶) in volume.

Cerebral cortex volumes of NOR and EAE mice were plotted against diseaseduration and a similar pattern was observed (FIG. 17B). Cerebral cortexvolumes were stable in the NOR group, while gradually decreasing in theEAE group (time×group interaction p=4.0×10⁻⁶). The volume of thecerebral cortex at sixty days after disease induction (d60) was 72.3 mm³(1.4 mm³) in NOR mice and 68.9 mm³ (1.8 mm) in EAE mice, a 4.7% decrease(p=1.5×10⁻¹) in volume.

Similarly, a progressive loss of cerebellar volume during EAE wasobserved in EAE mice compared to NOR mice (time×group interactionp=1.1×10⁻⁵)(FIG. 17C). The volume of the whole cerebellum at sixty daysafter disease induction (d60) was 53.2 mm³ (1.1 mm³) in NOR mice and50.0 mm (1.1 mm in EAE mice, a 6.0% decrease (p=3.1×10⁻⁵) in volume.

These results demonstrated that whole brain, cerebral cortex andcerebellum volume decreases over time in mice with EAE.

AC-186 EA vs. Vehicle EAE. A decrease in the rate of atrophy wasobserved in AC-186 treated mice compared to vehicle treated EAE micewhen whole brain volumes were plotted against the disease duration(starting with the scans prior to disease induction) (FIG. 17A). Inorder to quantify the effect of AC-186 treatment on EAE mice, arepeated-measures ANOVA was performed. Brain volume decreased in bothEAE and AC-186 mice, however this brain volume decrease wassignificantly smaller in AC-186 treated EAE mice (time×group interactionp=0.0013). The volume of the whole brain sixty days after diseaseinduction (d60) was 483 mm³ (7.0 mm³) in AC-186 treated EAE mice and 478mm³ (3.7 mm³) in vehicle treated EAE mice, indicating a 2.7% difference(p=0.021) in volume.

Cerebral cortex volumes of AC-186 treated EAE mice and vehicle treatedEAE mice were plotted against disease duration and a similar pattern wasobserved (FIG. 17B). Cerebral cortex atrophy rates were decreased inAC-186 treated EAE mice compared to vehicle treated EAE mice (time×groupinteraction p=0.0035). The volume of the cerebral cortex at sixty daysafter disease induction (d60) was 70.8 mm (1.6 mm³) in AC-186 treatedEAE mice and 68.9 mm³ (1.8 mm³) in vehicle treated EAE mice, a 5.0%difference (p=0.014) in volume.

When cerebellar volume was plotted against disease duration in AC-186treated EAE mice and vehicle treated EAE mice, volume decreases overtime were again observed (FIG. 17C). However, AC-186 treatment decreasedthe amount of volume loss compared to vehicle treatment (time×groupinteraction p−9.5×10). The volume of the whole cerebellum at sixty daysafter disease induction (d60) was 51.8 mm (1.1 mm³) in AC-186 treatedEAE mice and 50.0 mm (1.1 mm) in vehicle treated EAE mice, a 3.5%difference (p=0.0013) in volume.

Cerebral and cerebellar neuropathology revealed that AC-186 treatmentprevented neuronal cell (NcuN) and synaptic (PSD-95) loss in cerebralcortex gray matter (FIG. 18 top panel) and Purkinje neuronal cell(Calbindin) and synaptic (PSD-95) loss in the cerebellar cortex graymatter (FIG. 18 bottom panel).

Spinal Cord Neuropathy. Neuropathology was repeated on spinal cord toconfirm the results observed in Example 8. The results depicted in FIGS.19 and 20 largely reproduce the results in FIGS. 14 and 15, showing aprotective effect on axons. However, a beneficial effect of AC-186 ascompared to vehicle with regard to increased MBP staining, that wasobserved as a trend in Example 8 (FIG. 14), was observed as significant(FIG. 19). Together, these data indicate that given a large enoughsample size, AC-186 treatment does indeed have a beneficial effect onpreserving myelin staining in the CNS. Thus, AC-186 treatment on EAEresulted in sparing of axons and myelin in spinal cords. Notably, theeffect of AC-186 on lowering CD45 and Iba-1, but not CD3, which wasobserved in Example 8, was confirmed (FIG. 20). Lower CD45 and Iba-1staining in the CNS during AC-186 treatment is consistent with abeneficial effect of AC-186 treatment on clinical scores since thesecells are thought to contribute to the pathogenesis of disease.

Cerebellar and Cerebral White Matter Neuropathy. As an extension ofeffects of AC-186 treatment on spinal cord white matter neuropathology,AC-186 treatment on cerebral and cerebellar white matter was assessed.AC-186 administered at 30 mg/kg/every other day had a protective effecton cerebellar and cerebral axons (NF200) and myelin (MBP) (FIGS. 21 and22). These findings were consistent with the finding that AC-186protected spinal cord white matter. Together, these data show abeneficial effect for AC-186 treatment initiated after disease onset,which reduced myelin and axonal loss in the white matter of thecerebellum and cerebrum during EAE.

Cerebellar and Cerebral Gray Matter Neuropathy. As an extension ofeffects of AC-186 treatment on cerebellar and cerebral white matterneuropathology, the effects of AC-186 treatment on cerebral andcerebellar gray matter was assessed. As shown in FIG. 23, AC-186displayed a protective effect at a dose of 30 mg/kg/every other day forcerebellar cells (Calbindin+Purkinje cells) and synapses (PSD-95 andSynapsin 1). Further, AC-186 displayed a protective effect at a dose ofAC-186 30 mg/kg/every other day for cerebral cells (NeuN+ neurons) andsynapses (PSD-95) (FIG. 24). No effect was observed for Synapsin 1 incerebral gray matter, in contrast to cerebellar gray matter. Theseresults are consistent with the previous findings on the differentialeffect of EAE on pre (Synapsin 1) versus post (PSD-95) synaptic proteinexpression (Du et al., Proceedings of the National Academy of SciencesUSA, 111:2806-2806, 2014). Together, these data show a beneficial effectof AC-186 treatment, initiated after disease onset, in halting cellularand synaptic loss in cerebellar and cerebral gray matter during EAE.

Example 10

Examination of the Effect of Estrogen Receptor β Ligands, KBRV1 andKBRV2 on Neurodegeneration in the EAE Model.

Methods. Animals: Female WT C57BL/6 mice age 6-8 weeks at the time ofdisease onset, were obtained from Jackson Laboratories (Bar Harbor, Me.USA). Animals were maintained in accordance with the guidelines set bythe National Institute of Health and as mandated by the University ofCalifornia Los Angeles Office for the Protection of Research Subjects,the Chancellor's Animal Research Committee, and the PHS Policy on HumanCare and Use of Laboratory Animals.

Reagents: Two estrogen receptor β ligands, KBRV1 and KBRV2, wereprovided by Karo Bio (Huddinge, Sweden). Diarylpropionitrile (DPN) wasused as a positive control during the experiment. MOG peptide, aminoacids 35-55, was synthesized to >98% purity by Mimotopes (Clayton,Victoria, Australia).

Induction of EAE: In order to model brain inflammation, EAE was inducedin the mice. First, the mice were immunized by subcutaneous injectionsinto the left flank of 200 μg of MOG peptide, amino acids 35-55, and 200μg of Mycobacterium tuberculosis in complete Freund's adjuvant.Immediately after immunization, mice received an intraperitonalinjection of 500 ng pertussis toxin dissolved in 400 μL phosphatebuffered saline (PBS). Two days later, the mice received anotherintraperitoncal injection of pertussis toxin of the same quantity. Sevendays after the initial immunization, the MOG immunization was repeated.

Treatment: The mice were treated with vehicle, KBRV1, KBRV2, or DPNseven days prior to disease induction, Treatment was administered everyother day. DPN was administered as a positive control at a dose of 8mg/kg. Doses of KBRV1 included 3 μmol/kg and 10 μmol/kg. Doses of KBRV2were administered at 1 μmol/kg, 3 μmol/kg, and 101 μmol/kg.

Clinical Scoring: Animals were monitored daily for EAE signs based on astandard EAE 0 to 5 scale scoring system: 0, healthy; 1, complete lossof tail tonicity; 2, loss of righting reflex; 3, partial paralysis; 4,complete paralysis of one or both hind limbs; and 5, moribund.

Tissue Collection: In order to analyze the drug exposure of each animal,the plasma, brains, and spleen of the animals were taken from eachdosing group in Trial 1 (except for the group that were administeredDPN), as well as all groups in Trial 2, for bioanalysis. In Trial 3, thetissue was collected for histopathology.

Statistical Analysis: The results and measurements were evaluated usinga repeated measure ANOVA model with Bonferonni correction. Thestatistical significance level was set at 0.05, so any calculatedp-value less than 0.05 was considered significant.

Results. Trial 1: Trial 1 tested KBRV1 doses of 3 μmol/kg and 10μmol/kg, as well as KBRV2 doses of 10 μmol/kg. The results demonstratedthat both KBRV1 and KBRV2 were effective at ameliorating EAE whencompared to the vehicle treatment. The KBRV1 dose of 3 μmol/kg waseffective but the 10 μmol/kg dose was not effective. In performing thestatistical analysis, the F-test statistic of the 3 μmol/kg dosecompared to the vehicle was 36.59, giving a p-value less than 0.0001.The KBRV2 dose of 10 mol/kg was also effective when compared to thevehicle, giving an F-test statistic of 36.59 and a p-value of less than0.0001.

Trial 2: Trial 2 tested alternative doses of KBRV2 at both the 3 mol/kgand 10 μmol/kg level. The results demonstrated that KBRV2 was againeffective in ameliorating EAE when compared to the vehicle treatment.The 3 μmol/kg dose resulted in an F-test statistic of 29.20 and ap-value of less than 0.0001 when compared to the vehicle treatment.

Trial 3: Trial 3 repeated the tests of Trial 2 to confirm whether boththe 3 μmol/kg and 10 mol/kg doses of KBRV2 were successful inameliorating EAE. The results demonstrated that both doses wereeffective in ameliorating EAE. When comparing either dose to the vehicletreatments, the F-test statistic was calculated as 30.67, giving ap-value of less than 0.0001.

Treatment with FR ligand KBRV2 reduced demyelination and preservedmyelin in spinal cords of mice with EAE in a dose dependent manner withgreatest effects at the 10 μmol/kg dose and lesser effects at the 3μmol/kg dose. Treatment with ERβ ligand KBRV2 reduced axonal loss inspinal cords of mice with EAE in a dose dependent manner with greatesteffects at the 10 μmol/kg dose and lesser effects at the 3 μmol/kg dose.Further, treatment with ERβ ligand KBRV2 reduced local CNS inflammationand assessed by the global marker CD45 which stains all immune cells.This effect of ERβ ligand KBRV2 on inflammation within the CNS appearsto be due to effects on T lymphocytes, and not macrophages.

Example 11

Ovariectomized mice (OVX-Placebo) display worse performance in theMorris water maze than control mice (Sham-Placebo) (FIG. 19, top panel).The administration of an estrogen improves performance forovariectomized mice (OVX-Estriol) (FIG. 19, top panel). This effect islikely mediated by estrogen receptor β expressed by neurons becauseestrogen-mediated improvement is lost in mice comprising a conditionalknockout of estrogen receptor β in a neuron-specific enolase-CRE mouse(NSE-Crc:ERβ^(fl/fl); FIG. 19, bottom panel). Thus, estrogen receptor βligands can likely protect against neuronal defects and disabilityduring neurogenerative processes.

EQUIVALENTS

Although the present invention has been described in terms of theembodiments above, numerous modifications and/or additions to theabove-described embodiments would be readily apparent to one skilled inthe art.

INCORPORATION BY REFERENCE

All of the patents, published patent applications, and other documentscited herein are hereby incorporated by reference.

1-85. (canceled)
 86. A method of: i) slowing, reversing or haltingprogression of disability in a subject suffering from aneurodegenerative disease; ii) slowing or halting the progression ofmemory loss in a subject suffering from a neurodegenerative disease; oriii) slowing or halting the progression of a learning disability in asubject suffering from a neurodegenerative disease; wherein the methodcomprises administering an estrogen receptor beta ligand to the subject.87. The method of claim 86, wherein the method is a method of slowing,reversing or halting progression of disability in a subject sufferingfrom a neurodegenerative disease.
 88. The method of claim 86, whereinthe method is a method of slowing or halting the progression of memoryloss in a subject suffering from a neurodegenerative disease.
 89. Themethod of claim 86, wherein the method is a method of slowing or haltingthe progression of a learning disability in a subject suffering from aneurodegenerative disease.
 90. The method of claim 86, wherein thesubject has brain gray matter loss.
 91. The method of claim 90, whereinthe subject has greater than about 0.1% brain gray matter loss perannum.
 92. The method of claim 90, wherein the brain gray matter lossper annum is about 0.2%, or about 0.3%, or about 0.4%, or about 0.5%, orabout 0.6%, or about 0.7%, or about 0.8%, or about 0.9%, or about 1.0%,or about 1.1%, or about 1.2%, or about 1.3%, or about 1.4%, or about1.5%, or about 2.0%.
 93. The method of claim 90, wherein the gray matterloss is located in one or more of the total brain, cerebral cortex,cerebellum, thalamus, caudate nucleus, or putamen.
 94. The method ofclaim 90, wherein the gray matter loss is located in the cerebralcortex.
 95. The method of claim 90, wherein the estrogen receptor betaligand slows or eliminates the gray matter loss.
 96. The method of claim90, wherein the gray matter loss is measured using an imaging techniqueor surrogate marker.
 97. The method of claim 90, wherein the loss ofbrain gray matter is measured by MRI.
 98. The method of claim 86,wherein the estrogen receptor beta ligand is administered on a dailybasis.
 99. The method of claim 86, wherein the neurodegenerative diseaseis Alzheimer's disease, Parkinson's disease, multiple sclerosis, stroke,amyotrophic lateral sclerosis, cerebellar ataxia, frontotemporaldementia, prion disease, Huntington's disease, cerebral ischemia,cerebral dementia syndrome, infection-induced neurodegenerationdisorders, AIDS-encephalopathy, Creutzfeld-Jakob disease,encephalopathies induced by rubiola, herpes viruses and borrelioses,metabolic-toxic neurodegenerative disorders, encephalopathies induced bysolvents or pharmaceuticals, trauma-induced brain damage, or spinal cordinjury.
 100. The method of claim 86, wherein the neurodegenerativedisease is Alzheimer's disease, Parkinson's disease, trauma-inducedbrain damage, or spinal cord injury.
 101. The method of claim 86,wherein the estrogen receptor beta ligand is conjointly administeredwith an immunotherapeutic agent.
 102. The method of claim 86, whereinthe immunotherapeutic agent is interferon-beta 1a, interferon-beta 1b,pegylated interferon-beta-1a, glatiramer acetate, natalizumab,mitoxantrone, fingolimod, teriflunomide, dimethyl fumarate,mycophenolate mofetil, paclitaxel, cyclosporine, corticosteroids,azathioprine, cyclophosphamide, methotrexate, cladribine,4-aminopyridine, and tizanidine.
 103. The method of claim 86, whereinthe subject is female.
 104. The method of claim 86, wherein the methodis a method of slowing or halting the progression of memory loss in aneurodegenerative disease subject; the subject has brain gray matterloss; the subject has mild cognitive impairment; and the subject isfemale.
 105. The method of claim 86, wherein the method is a method ofslowing or halting the progression of memory loss in a neurodegenerativedisease subject; the subject has brain gray matter loss; the gray matterloss is located in the cerebral cortex; the neurodegenerative disease isAlzheimer's disease; and the subject is female.